Identical Pulsed-field Gel Electrophoresis Research Articles

A Community Waterborne Salmonella Bovismorbificans Outbreak in Greece.

In August 2022, the Hellenic National Public Health Organisation was notified about a gastroenteritis outbreak in town A in Southern Greece. Investigations aimed to identify the source and implement control measures. Case definition categories were used in a 1:3 case-control study. Cases and controls were interviewed about various exposures. Cases' stool samples were cultured on agar plates and characterised by serotyping, antimicrobial susceptibility testing and Pulse Field Gel Electrophoresis (PFGE). Environmental investigations included tap water sampling for microbiological and chemical analysis in town A and inspection of the water supply system. We identified 33 cases (median age: 17 years). Tap water consumption was the only significant risk factor for gastroenteritis (OR = 5.46, 95% CI = 1.02-53.95). Salmonella (S.) Bovismorbificans isolated from eight stool and one tap water samples had identical PFGE profiles. No resistant isolates were identified. Residual chlorine levels were lower than the acceptable limits before and during the outbreak. We advised consumption of bottled water and adherence to strict hand hygiene rules until tap water was declared suitable for drinking. Epidemiological and molecular data revealed a waterborne S. Bovismorbificans outbreak in town A. We recommend local water safety authorities to ensure that residual chlorine levels comply with the legislation towards water safety planning, to mitigate risks.

Different cellular and molecular responses of Bovine milk phagocytes to persistent and transient strains of Streptococcus uberis causing mastitis.

Streptococcus uberis is frequently isolated from milk collected from dairy cows with mastitis. According to the host's immunity, bacterial virulence, and their interaction, infection with some strains can induce persistent subclinical inflammation, while infection with others induces severe inflammation and transient mastitis. This study compared the inflammatory response of milk-isolated white blood cells (mWBCs) to persistent and transient S. uberis strains. Quarter milk samples were collected aseptically for bacterial culture from all lactating cows once a week over a 10-week period. A transient and noncapsular strain with a 1-week intramammary infection duration was selected from this herd, while a persistent and capsular S. uberis strain with an intramammary infection longer than 2 months from our previous study was selected based on an identical pulse field gel electrophoresis pattern during the IMI episode. Cellular and molecular responses of mWBCs were tested, and the data were analyzed using repeated analysis of variance. The results showed a higher response in migration, reactive oxygen species generation, and bacterial killing when cells were stimulated with transient S. uberis. In contrast, the persistent strain led to increased neutrophil extracellular trap release. This study also highlighted several important molecular aspects of mWBCs. Gene expression analyses by real-time RT-PCR revealed a significant elevation in the expression of Toll-like receptors (TLR-1, TLR-2, TLR-6) and proinflammatory cytokines (tumor necrosis factor-alpha or TNF-α) with the transient strain. Additionally, Streptococcus uberis capsule formation might contribute to the capability of these strains to induce different immune responses. Altogether, these results focus on the immune function of activated mWBCs which demonstrate that a transient strain can elicit a stronger local immune response and, subsequently, lead to rapid recovery from mastitis.

Epidemiological and molecular evidence of foodborne poisoning outbreak caused by enterotoxin gene cluster-harboring Staphylococcus aureus of new sequence type 7591

ObjectivesThis paper presented a detailed analysis of the epidemiology and molecular characteristics of staphylococcal food poisoning (SFP) that occurred in a hotel in Hangzhou. MethodsA total of 46 guests at the hotel underwent an epidemiological survey. Samples of stool from patients, vomit, swabs from the kitchen, leftover food items, and anal swabs from food handlers were taken and investigated for the presence of potential pathogenic bacteria. Molecular techniques and whole genome sequencing were performed to track the evolution of Staphylococcus aureus associated with the outbreak of SFP. ResultsForty-six individuals displayed gastrointestinal symptoms. Seventeen isolates of S. aureus were discovered to carry the seg, sei, sem, sen, seo, and selu genes found in a specific enterotoxin gene cluster (egc) operon, but without the presence of classical enterotoxins such as SEA ∼ SEE. All egc-positive isolates shared identical pulsed-field gel electrophoresis profiles and were classified under new ST7591 (Clonal Complex 72) with identical spa typing t148. In addition, some isolates of S. aureus obtained from food sources sold in Hangzhou over the past 3 years and carrying egc genes were grouped under the ST72 lineage (CC72). Through whole genome sequencing, a strong genetic connection was revealed between these egc-positive isolates and clinical ST72 S. aureus found in China. ConclusionsS. aureus with non-classical egc enterotoxins was suggested to be a potential cause of SFP in humans.

Antimicrobial resistance and molecular epidemiological analysis of Escherichia fergusonii harboring the mcr gene in pigs and broiler chickens in Okinawa, Japan.

The dissemination of mcr-harboring Enterobacteriaceae, e.g., Escherichia fergusonii, with resistance to colistin via animal products is a public health concern. In our previous study, E. fergusonii harboring the mcr gene were isolated from 11 pigs and 43 chickens. To understand the spread of mcr-harboring E. fergusonii in Okinawa, Japan, and to gain further insights into how they can be controlled, an antimicrobial susceptibility testing, pulsed-field gel electrophoresis (PFGE), a conjugation test for the transferability of mcr-harboring plasmids, and PCR-based replicon typing (PBRT) were performed using the 54 strains. According to the disk-diffusion and broth microdilution methods, 9 of the 11 strains from pigs and 9 of the 43 strains from chickens had multidrug resistance (MDR). The broth microdilution method showed that all strains were resistant to colistin, and the minimum inhibitory concentration of colistin was 4-16 μg/mL. PFGE suggested identical PFGE types were being transmitted within one pig farm, within one chicken farm, and among several chicken farms. These findings showed that some mcr-harboring E. fergusonii in Okinawa exhibited MDR, and these had spread within farms and between farms. In the mcr gene conjugation test and PBRT, a type IncI2 plasmid replicon was detected in all mcr-1-harboring transconjugants. Therefore, evidence suggests that the IncI2 plasmid is probably involved in the transmission of the mcr-1 gene. It is important to monitor the antimicrobial resistance profile and dissemination of the IncI2 plasmid in mcr-harboring E. fergusonii.

Mitigation strategies to control a carbapenem-resistant Acinetobacter baumannii outbreak in a dedicated COVID-19 unit

Background: Carbapenem-resistant Acinetobacter baumannii (CRAb) is considered a public health threat, and this pathogen is typically associated with hospital infections. At 3 months after opening the hospital’s dedicated COVID-19 unit, 2 patients were identified with CRAb. Infection prevention staff collaborated with staff in the COVID-19 unit, hospital leadership, and health department partners to develop mitigation strategies and to prevent additional transmission. Methods: Admissions to the COVID-19 unit were stopped. Biweekly surveillance cultures were collected to identify any patients potentially colonized with CRAb. An infection control risk assessment was conducted to determine breaches in infection prevention practices. The risk assessment included environmental rounding of the area, epidemiological investigation, environmental testing, pulsed-field gel electrophoresis (PFGE) testing, and observing infection prevention practices. Results: The risk assessment identified multiple gaps in infection control practices, for example, gaps in hand and environmental hygiene practices. The extended use of personal protective equipment (PPE), staff shortages, fatigue, and staff taking on multiple roles and tasks outside their general job duties were other gaps identified by the team. Between June and September 2020, 43 additional CRAb cases were identified in the facility, with 4 (9.8%) cases outside the COVID-19 unit. Moreover, 29 cases (64%) were considered clinical infections and 16 (36%) were identified from surveillance efforts. Environmental cultures identified 1 positive surface with CRAb. PFGE testing was completed on 44 isolates; 42 isolates had identical PFGE patterns, and 2 isolates were unrelated to the COVID-19 unit; 2 isolates were closely related (with 3-band differences) but were not identified in the COVID-19 unit. Conclusions: The inability to definitively identify the source of transmission made it challenging to determine the best approach to eradicating the pathogen. Mitigation for outbreaks should focus on not deviating from core infection control practices.Funding: NoneDisclosures: None

Early identification of a clustered epidemic of typhoid fever through pathogen molecular subtyping monitoring

Objective: To identify a suspected clustered Typhoid fever by whole genome sequencing(WGS) and pulsed field gel electrophoresis (PFGE) subtyping. Methods: The nature of the epidemic was determined by combination of subtyping results of isolates and epidemiological information. Results: Five S. typhimurium isolates showed identical PFGE patterns and almost the same whole genome sequence. Epidemiological survey showed that five cases had dined in the same restaurant on the same day. Conclusion: Combined with the longest incubation period of typhoid fever, molecular subtyping of pathogenic bacteria and the field epidemiological survey, it can be preliminarily determined that the five cases have common infection sources.

Genomic Investigation Reveals a Community Typhoid Outbreak Caused by Contaminated Drinking Water in China, 2016

Typhoid fever is a life-threatening disease caused by Salmonella enterica serovar Typhi (S. Typhi) and remains a significant public health burden in developing countries. In China, typhoid fever is endemic with a limited number of reported outbreaks. Recently, Chinese local Center for Disease Prevention and Control is starting to apply whole genome sequencing for tracking the source of outbreak isolates. In this study, we conducted a retrospective investigation into a community outbreak of typhoid fever in Lanling, China, in 2016. A total of 26 S. Typhi isolates were recovered from the drinking water (n = 1) and patients' blood (n = 24) and stool (n = 1). Phylogenetic analysis indicated the persistence of the outbreak isolates in drinking water for more than 3 months. The genomic comparison demonstrated a high similarity between the isolate from water and isolates from patients in their genomic content, virulence gene profiles, and antimicrobial resistance gene profile, indicating the S. Typhi isolate from drinking water was responsible for the examined outbreak. The result of pulsed-field gel electrophoresis (PFGE) revealed these isolates had identical PFGE pattern, indicating they are clonal variants. Additionally, phylogeographical analysis of global S. Typhi isolates suggested the outbreak isolates are evolutionarily linked to the isolates from the United Kingdom and Vietnam. Taken together, this study highlights the drinking water and international travel as critical control points of mitigating the outbreak, emphasizing the necessity of regular monitoring of this pathogen in China.

Prevalence of qnrS-positive Escherichia coli from chicken in Thailand and possible co-selection of isolates with plasmids carrying qnrS and trimethoprim-resistance genes under farm use of trimethoprim

One hundred and twenty chicken samples from feces (n = 80), the carcass surface at slaughter at 2 meat chicken farms (n = 20), and retail chicken meat from 5 markets (n = 20) collected during 2018 and 2019 were examined for the prevalence of plasmid-mediated quinolone resistance (PMQR) in Escherichia coli. We detected qnrS-positive E. coli in a total of 74 samples from feces (n = 59), the carcass surface (n = 7), and retail meat (n = 8). These 74 qnrS-positive isolates were tested for antimicrobial susceptibility to determine the minimum inhibitory concentrations (MICs) of certain antimicrobials and genetically characterized. Ampicillin-resistance accounted for 71 of the 74 isolates (96%), followed by resistance to oxytetracycline (57/74; 77%), enrofloxacin (ERFX) (56/74; 76%), sulfisoxazole (SUL) (56/74; 76%), trimethoprim (TMP) (49/74; 66%), and dihydrostreptomycin (48/74; 65%). All farm-borne SUL- and TMP-resistant isolates except one were obtained from samples from farm A where a combination of sulfadiazine and TMP was administered to the chickens. Concentrations of ERFX at which 50 and 90% of isolates were inhibited were 2 μg/mL and 32 μg/mL, respectively. Diverse pulsed-field gel electrophoresis (PFGE) patterns of XbaI-digested genomic DNA were observed in the qnrS-positive isolates from fecal samples. Several isolates from feces and the carcass surface had identical XbaI-digested PFGE patterns. S1-nuclease PFGE and Southern blot analysis demonstrated that 7 of 11 dfrA13-positive fecal isolates carried both the qnrS and dfrA13 genes on the same plasmid, and 2 of 3 dfrA1-positive isolates similarly carried both qnrS and dfrA1 on the same plasmid, although the PFGE patterns of XbaI-digested genomic DNA of the isolates were different. These results suggest that the qnrS gene is prevalent in chicken farms via horizontal transfer of plasmids and may partly be co-selected under the use of TMP.

Genetic Characterization of CTX-M-2-Producing Klebsiella pneumoniae and Klebsiella oxytoca Associated With Bovine Mastitis in Japan.

CTX-M-2-producing Klebsiella oxytoca (K. oxytoca) has not received much attention in animal husbandry compared with Klebsiella pneumoniae (K. pneumoniae), a major reservoir of extended-spectrum β-lactamase (ESBL) genes. Bacteriological examinations of 1,466 mastitic milk samples between October 2012 and December 2014 were conducted. Ninety-five K. pneumoniae isolates (total prevalence: 6.5%) and 81 K. oxytoca isolates (total prevalence: 5.5%) were obtained. Seventeen K. pneumoniae isolates obtained from 15 animals reared on 11 farms and 9 K. oxytoca isolates obtained from 9 animals reared on the same farm were phenotypically confirmed to be ESBL producers. All nine ESBL-producing K. oxytoca isolates were obtained from one farm between June and November 2013 and related to a significantly (p < 0.05) higher monthly prevalence of mild mastitis (in June, August, September, October, and November 2013). Pulsed-field gel electrophoresis (PFGE) patterns of ESBL-producing K. pneumoniae isolates were distinguished from each other by more than 6-band differences except for two isolates from two animals, whereas all nine K. oxytoca isolates showed an identical PFGE pattern. Transferability of the blaCTX−M−2 gene was found in 14 K. pneumoniae and 9 K. oxytoca isolates by conjugation analysis. Of these isolates, the blaCTX−M−2 gene was detected on plasmids belonging to the incompatibility (Inc) groups P and N derived from five K. pneumoniae and nine K. oxytoca isolates, respectively, although the plasmids from the remaining nine K. pneumoniae were untypeable. All the transconjugants exhibited elevated minimum inhibitory concentrations of ampicillin, cefotaxime, and ceftiofur compared with those in the wild-type, recipient strain. Restriction fragment length polymorphism analysis demonstrated that the IncN plasmids extracted from eight of nine transconjugants, which received resistance against β-lactams from K. oxytoca, showed an identical DraI digestion pattern. These results suggest that the CTX-M-2-producing K. oxytoca strain with the above-mentioned characteristics may have clonally spread within a farm, whereas the blaCTX−M−2 gene in K. pneumoniae possibly disseminated among the farms through different plasmids. Thus, monitoring of ESBL genes, including the blaCTX−M−2 gene, among causative agents of bacterial mastitis in cows can help to develop relevant treatments and control practices.

Clinical and microbiological characteristics of nosocomial, healthcare-associated, and community-acquired Klebsiella pneumoniae infections in Guangzhou, China

BackgroundKlebsiella pneumoniae (K. pneumoniae) is a common pathogen associated with hospital and community-onset infections. This study aimed to compare the clinical and microbiological characteristics of nosocomial, healthcare-associated (HCA), and community-acquired (CA) K. pneumoniae infections.MethodsClinical data were extracted from electronic medical records and analyzed retrospectively. Antimicrobial susceptibility and extended-spectrum beta-lactamase (ESBL) production were determined for all identified strains. Carbapenemase and ESBL genes were amplified by PCR. Genotyping of carbapenem-resistant K. pneumoniae (CRKP) and ESBL-producing strains was performed by pulsed-field gel electrophoresis (PFGE).ResultsOf 379 K. pneumoniae infections, 98 (25.9%) were nosocomial, 195 (51.5%) were healthcare-associated, and 86 (22.6%) were community-acquired. Hematological malignancy (OR = 4.467), and hypertension (OR = 2.08) and cerebral vascular disease (OR = 2.486) were associated with nosocomial and HCA infections respectively, when compared to CA infections. Overall, the incidence of antimicrobial resistance for the majority of agents tested was similar between nosocomial and HCA infections (P > 0.05) and both groups had a higher incidence than CA infections (P < 0.05). Moreover, 95.1% (78/82) of CRKP strains were isolated from the nosocomial and HCA groups. The blaKPC was the most prevalent carbapenemase gene among CRKP strains (80.5%, 66/82). ESBL-producing strains were prevalent among nosocomial (40.8%), HCA (35.9%) and CA groups (24.4%). The blaCTX-M-9-group and blaCTX-M-1-group genes were predominant in nosocomial (65.0%) and CA strains (66.7%), respectively. PFGE results showed ESBL-producing and CRKP strains were genetically diverse. Identical PFGE profiles were observed among HCA and nosocomial strains.ConclusionsNosocomial and HCA K. pneumoniae infections presented similar clinical features and antimicrobial resistance, and both two types of infections were different to CA infections. CRKP and ESBL-producing strains were disseminated mainly in HCA and nosocomial groups, and showed a clonal diversity. The cross transmission of CRKP was existed among HCA and nosocomial patients. This finding suggests that similar empirical therapy should be considered for patients with nosocomial and HCA K. pneumoniae infections and bacterial resistance surveillance of these infections is necessary.

Efficacy of a coordinated strategy for containment of multidrug-resistant Gram-negative bacteria carriage in a Neonatal Intensive Care Unit in the context of an active surveillance program

BackgroundAntimicrobial resistance in neonatal intensive care unit (NICU) patients is a threat, due to the frequent use of antimicrobial treatment and invasive devices in fragile babies. Since 2014 an active surveillance program of multidrug-resistant Gram-negative bacteria (MDR-GNB) carriage has been in place in the five NICUs of Palermo, Italy. In 2017 an increase in the prevalence of MDR-GNB, and in particular of extended-spectrum β-lactamases-producing Klebsiella pneumoniae (ESBL-KP), was observed in “Civico” hospital NICU.AimTo assess the impact of a coordinated intervention strategy in achieving long-lasting reduction of MDR-GNB prevalence in the NICU.MethodsRectal swabs were obtained monthly and processed to detect MDR-GNB using standard methods. MDR-GNB were characterized by pulsed-field gel electrophoresis (PFGE). Since November 2017 the following intervention measures were applied: (a) two-months intensification of sample collection; (b) stakeholders meetings; (c) improvement of prevention measures and antimicrobial policies.FindingsDuring the intensified microbiological surveillance MDR-GNB and ESBL-KP were detected in rectal swabs (34.8%; 23.2%), nasal swabs (24.6%; 14.5%), oral swabs (14.5%; 5.4%), milk samples (32.1%; 17.9%), pacifiers swabs (30.8%; 17.9%) and from sub-intensive room surfaces. Thirteen ESBL-KP strains isolated from clinical and environmental samples showed identical PFGE patterns. The prevalence of MDR-GNB and ESBL-KP carriage significantly decreased in the year after intervention compared to the previous year (20.6% vs 62.2%; p < 0.001 and 11.1% vs 57.8%; p < 0.001). MDR-GNB were not detected at all for three months and ESBL-KP for five months. Multivariate analysis of the principal exposure variables showed that admission in the post-intervention period significantly reduced the risk of MDR-GNB carriage (adj-OR = 0.21, 95% CI = 0.076–0.629; p < 0.001).ConclusionsMDR-GNB broadly circulate in NICU setting, they can colonize different body sites and spread through various vehicles. A coordinated strategy of multiple interventions with active cooperation between epidemiologists and clinicians in the NICU can effectively reduce their circulation and in particular the carriage of the most dangerous ESBL-KP strains.

Emergence of blaNDM-5 in Enterobacteriaceae Isolates from Companion Animals in Guangzhou, China.

The occurrence and characterization of carbapenemase-producing Enterobacteriaceae from companion animals in Guangzhou, China, are investigated. Six isolates (2.3%, 6/257) were positive for blaNDM-5, that is, one Enterobacter cloacae, one Citrobacter freundii, and four Escherichia coli. Three E. coli isolates obtained from the same animal hospital were ST410 and showed identical pulse field gel electrophoresis pattern, resistance profiles, and resistance genes. blaNDM-5 was located on IncX3 (n = 5) and IncK2 (n = 1) plasmid, respectively. The presence of carbapenemase-producing Enterobacteriaceae among companion animals needs continued surveillance.

Family Clusters of Shiga Toxin-producing Escherichia coli Infection: An Overlooked Source of Transmission. Data From the ItalKid-Hus Network.

The aim of the present work was to investigate family clusters of Shiga toxin-producing Escherichia coli (STEC) infection among the household members of STEC positive patients, identified within a screening program of bloody diarrhea (BD) for STEC in Northern Italy. Stool samples from patients with BD or BD-associated-hemolytic uremic syndrome (HUS) and related households were investigated by molecular and bacteriologic methods to detect and characterize the virulence profile of STEC and Pulsed Field Gel Electrophoresis analysis were done on isolates. Thirty-nine cases of STEC infection (isolated BD in 16, BD-associated-HUS in 23) were considered, and a total of 130 stool samples from 1 to 8 households of the index patient were analyzed. The prevalence of positivity was higher in siblings (34.8%, 8/23) than in mothers (20%, 7/35), grandparents (9.5%, 2/21), fathers (8.8%, 3/34) or other households. In 14 clusters (36%), one or more household shared a STEC with the same virulence profile (stx, eae, serogroup) as the index case. In 7 clusters, STEC strains isolated from at least 2 subjects also shared identical Pulsed Field Gel Electrophoresis profile. The frequency of household infection does not appear to be associated to the index case's illness (HUS or BD), nor with the serotype or with the virulence profile of the involved STEC (stx2 or stx1-stx2). Our study shows that STEC infections, most likely related to human-to-human transmission, are common among households of patients with STEC BD or HUS and underlines the importance of extending the epidemiologic investigations to all family members, as the index case may not always be the primary infection in the family.

Coagulase-Negative Staphylococci Contained in Gut Microbiota as a Primary Source of Sepsis in Low- and Very Low Birth Weight Neonates

Background: There are only a few reports in the literature about translocation of coagulase-negative staphylococci (CoNS) as a primary cause of sepsis in neonates, although CoNS are among a short list of “translocating” bacteria when present in abundance. Methods: 468 blood samples, 119 stool samples, and 8 catheter tips, from 311 neonates, were tested for presence of microorganisms. CoNS strains isolated from the blood and stool or from blood and catheter tip of the same newborn at approximately the same time were paired and typed with PFGE (Pulse-Field Gel Electrophoresis) method. The strains were then tested for the presence of adherence genes and biofilm formation. Results: The strains with identical PFGE profiles in comparison to those with non-identical profiles differed in terms of the pattern of the virulence genes and showed a lack of the genes related to adherence, but more often presence of IS256, which is related to virulence. They also were phenotypically unable to adhere to intestinal Caco2 cells. Conclusions: A considerable proportion of CoNS strains isolated from bloodstream of VLBW/LWB neonates was identical to the strains isolated from faeces of the same neonates at the same time. These observations may offer indirect evidence indicating that at least some CoNS can translocate from the gastrointestinal tract of the premature neonates into the bloodstream and thus cause generalized infection.

Detection of spatial, temporal and space-time Salmonella Heidelberg and Salmonella Typhimurium clusters in Ontario in 2015, and comparisons to known outbreaks.

Salmonellosis is one of several reportable diseases in Ontario (ON). Two or more cases of the same serotype that are linked to a common exposure or related to one another in time and/or space are considered a potential outbreak. While laboratory data can help to determine the molecular relatedness of cases, results may take up to several weeks. This study aimed to assess the utility of the retrospective spatial scan statistic in detecting clusters of Salmonella Heidelberg and Salmonella Typhimurium cases using data from ON in 2015. Identified clusters were validated by laboratory data (where available) to determine whether identified clusters were likely outbreaks. Data representing the location of each reported S. Heidelberg or S. Typhimurium case in 2015, responsible serotype and symptom onset date were exported to SaTScan for retrospective spatial, temporal, and space-time analyses using the spatial scan statistic with Bernoulli models and a space-time permutation model. Analyses were performed with and without those cases linked to known outbreaks. Laboratory subtyping data (i.e. pulsed field gel electrophoresis (PFGE) and/or phage type) and food and environmental exposure information (e.g. travel, animal contact, poultry and other food item consumption) were used to explore the relatedness of cases within identified clusters. Spatial, temporal and space-time analyses identified a known outbreak of S. Heidelberg in 2015 (n=9 cases) and a previously unidentified cluster of S. Heidelberg cases. Most cases (94%) within a cluster detected via a space-time permutation model of S. Heidelberg cases shared an identical PFGE pattern and appeared to represent a true outbreak. The spatial scan statistic, and particularly the space-time permutation model, could assist in outbreak identification before laboratory data are available, allowing for faster cluster identification and implementation of control measures.

The Predominance of Strain Replacement Among Enterobacteriaceae Pairs With Emerging Carbapenem Resistance During Hospitalization.

Isolates of Enterobacteriaceae collected from the same patient can lose carbapenem susceptibility during antimicrobial therapy, but little attention has been given to how this conversion takes place. In the current study, we retrospectively analyzed microbiological and clinical data from patients with enterobacterial infections at a tertiary hospital in Shanghai, China. After screening 4795 patients and 7120 Enterobacteriaceae isolates over the 3-year study period, we found the change from carbapenem susceptible to carbapenem resistant in 41 pairs of isolates, of which 35 pairs (85.4%) were K. pneumoniae and 25 (61.0%) were from the same anatomic sites. Thirty-six isolate pairs showed different pulsed-field gel electrophoresis patterns between the carbapenem-susceptible and the corresponding resistant strain, and 5 pairs displayed identical pulsed-field gel electrophoresis patterns. Thirty-three (91.7%) of the 36 pairs of Enterobacteriaceae isolates were carbapenem-resistant K. pneumoniae with blaKPC-2, and 28 pairs (90.3%) of K. pneumoniae isolates had different sequence types (STs), with ST11 the most common ST found in carbapenem-resistant K. pneumoniae isolates. Forty of the 41 patients had received antimicrobial therapy such as carbapenems, cephalosporins, and fluoroquinolones, before the isolation of carbapenem-resistant Enterobacteriaceae. These results demonstrated that strain replacement is the main cause of emerging carbapenem resistance in Enterobacteriaceae during hospitalization. The loss of carbapenem susceptibility was not mainly due to in vivo development of carbapenem resistance.

Emergence of a clone of invasive fucK-negative serotype e Haemophilus influenzae in British Columbia.

Introduction of the Haemophilus influenzae serotype b (Hib) conjugate vaccine has changed the epidemiology of invasive H. influenzae disease, with most infections now caused by non-typeable (non-encapsulated) and non-Hib encapsulated strains. We describe nine invasive serotype e H. influenzae (Hie) from British Columbia that were determined to have complete deletion of their fucose operon genes. These nine isolates were recovered from blood cultures of three female and six male patients during 2011-2018, with eight recovered in the past 4 years. All nine isolates were biotype IV, with eight showing identical pulsed field gel electrophoresis (PFGE) profiles, whereas the ninth showed 95% similarity. PFGE analysis also showed these fucose operon-negative Hie to be most (94%) similar to the multi-locus sequence type (ST)-18, the most common ST among Hie in British Columbia. These nine fucose operon-negative Hie represented 27.3% of the 33 invasive Hie isolated in British Columbia from 2010 to 2018. Deletion of the fucose operon did not appear to impact the transmission ability of these strains or their ability to cause invasive disease.

Emergence of two novel sequence types (3366 and 3367) NDM-1- and OXA-48-co-producing K. pneumoniae in Italy.

The aim of this study was to analyze the alarming spread of NDM-1- and OXA-48-co-producing Klebsiella pneumoniae clinical isolates, collected between October 2016 and January 2018 in a neonatal intensive care unit of the University Hospital, Catania, Italy, through whole genome sequencing. All confirmed carbapenem-resistant K. pneumoniae (CRKp) isolates were characterized pheno- and geno-typically, as well as by whole genome sequencing (WGS). A total of 13 CRKp isolates were identified from 13 patients. Pulsed-field gel electrophoresis (PFGE) was performed, and the multilocus sequence typing (MLST) scheme used was based on the gene sequence as published on the MLST Pasteur website. Core genome MLST (cgMLST) was also performed. All isolates co-carried blaoxa-48 and blaNDM-1 genes located on different plasmids belonging to the IncM/L and IncA/C2 groups, respectively. The 13 strains had identical PFGE profiles. MLST and cgMLST showed that K. pneumoniae was dominated by CRKp ST101 and two novel STs (ST3666 and ST3367), identified after submission to the MLST database for ST assignment. All isolates shared the same virulence factors such as type 3 fimbriae, genes for yersiniabactin biosynthesis, yersiniabactin receptor, and iron ABC transporter. They carried the wzi137 variant associated with the K17 serotype. To the best of our knowledge, this is the first report of two novel STs, 3366 and 3367, NDM-OXA-48-co-producing K. pneumoniae clinical isolates, in Italy.

Detection and Molecular Characterization of Staphylococci from Eggs of Household Chickens.

Eggs are a healthy and nutritious food source, but may be contaminated by bacteria. Previous studies have reported the presence of staphylococci in eggs of farmed chickens, but no study has evaluated the staphylococcal population of eggs from household chickens. In this study, staphylococci from eggs (n = 275) of household chickens collected from November 2016 to March 2017 from different villages of Khyber Pakhtunkhwa province, Pakistan, were characterized. Seven species of staphylococci were identified from 65 eggs, including the predominant species, Staphylococcus xylosus (49/275; 17.8%). S. xylosus isolates (n = 73) were tested for antimicrobial susceptibility, presence of resistance genes, genetic relatedness, and inhibitory activity against other bacteria. The majority of isolates were resistant to oxacillin (83.6%) and tetracycline (24.7%), but also exhibited resistance to daptomycin and linezolid (5.5% each). Of the 10 resistance genes tested, isolates were only positive for mecA (35.6%; 26/73), mecC/C1 (2.7%; 2/73), and tet(K) (14/73; 19%). Using pulsed-field gel electrophoresis (PFGE), nine clusters had identical PFGE patterns. Isolates produced inhibitory activity against a broad spectrum of bacteria; 20.5%, 19.2%, 17.8%, and 16.4% of S. xylosus were able to inhibit growth of Salmonella enterica serotype Typhi, methicillin-susceptible Staphylococcus aureus, Escherichia coli, and methicillin-resistant Staphylococcus aureus, respectively. This study demonstrated the presence of genetically related antimicrobial-resistant S. xylosus from eggs from household chickens. Like table eggs, eggs of household chickens also contain staphylococci that may be resistant to antimicrobials used to treat human infections. These data will allow comparison between staphylococci from eggs from different sources and may indicate the relative safety of eggs from household chickens. Further study of these egg types and their microbial composition is warranted.

Pediatric oncology and stem cell transplant patients with healthcare-associated Clostridium difficile infection were already colonized on admission.

A prospective cohort study was conducted over a three-year period in an inpatient pediatric oncology and stem cell transplant setting. Baseline stool samples were collected within three days of admission and were genotypically compared with clinically indicated samples submitted after three days of admission. A total of 175 patients were recruited with a total of 536 admissions. The adjusted prevalence of baseline toxigenic C. difficile colonization among admissions was 32.8%. Seventy-eight percent of positive admissions did not have history of CDI. Colonization with a toxigenic strain on admission was predictive of CDI (OR=28.6; 95% CI, 6.58-124.39; P<0.001). Nearly all clinical isolates (8/9) shared identical pulsed-field gel electrophoresis patterns with baseline isolates or were closely related (1/9). Only one of the 11 cases that were considered HA-CDI was potentially nosocomially acquired. The prevalence of colonization with toxigenic C. difficile in our cohort is high. Unfortunately, the current CDI surveillance definitions overestimate the incidence of HA-CDI in pediatric oncology and stem cell transplantation settings.