Nitric Oxide Induced stx2 Expression Is Inhibited by the Nitric Oxide Reductase, NorV, in a Clade 8 Escherichia coli O157:H7 Outbreak Strain

Escherichia coli O157:H7 pathogenesis is due to Shiga toxin (Stx) production, though variation in virulence has been observed. Clade 8 strains, for instance, were shown to overproduce Stx and were more common among hemolytic uremic syndrome cases. One candidate gene, norV, which encodes a nitric oxide (NO) reductase found in a clade 8 O157:H7 outbreak strain (TW14359), was thought to impact virulence. Hence, we screened for norV in 303 O157 isolates representing multiple clades, examined stx2 expression following NO exposure in TW14359 for comparison to an isogenic mutant (ΔnorV), and evaluated survival in THP-1 derived macrophages. norV was intact in strains representing clades 6–9, whereas a 204 bp deletion was found in clades 2 and 3. During anaerobic growth, NO induced stx2 expression in TW14359. A similar increase in stx2 expression was observed for the ΔnorV mutant in anaerobiosis, though it was not impaired in its ability to survive within macrophages relative to TW14359. Altogether, these data suggest that NO enhances virulence by inducing Stx2 production in TW14359, and that toxin production is inhibited by NorV encoded by a gene found in most clade 8 strains. The mechanism linked to these responses, however, remains unclear and likely varies across genotypes.

Mycobacterium intracellulare subsp. chimaera from Cardio Surgery Heating-Cooling Units and from Clinical Samples in Israel Are Genetically Unrelated

Non-tuberculous mycobacteria (NTM) are opportunistic pathogens that cause illness primarily in the elderly, in the immunocompromised or in patients with underlying lung disease. Since 2013, a global outbreak of NTM infection related to heater-cooler units (HCU) used in cardio-thoracic surgery has been identified. This outbreak was caused by a single strain of Mycobacterium intracellulare subsp. chimaera. In order to estimate the prevalence of this outbreak strain in Israel, we sampled Mycobacterium intracellulare subsp. chimaera from several HCU machines in Israel, as well as from patients, sequenced their genomes and compared them to the outbreak strain. The presence of mixed mycobacteria species in the samples complicated the analysis of obtained sequences. By applying a metagenomic binning strategy, we were able to obtain, and characterize, genomes of single strains from the mixed samples. Mycobacterium intracellulare subsp. chimaera strains were compared to each other and to previously reported genomes from other countries. The strain causing the outbreak related to the HCU machines was identified in several such machines in Israel but not in any clinical sample.

Mycobacterium chimaera infections among cardiothoracic surgery patients associated with heater-cooler devices—Kansas and California, 2019

Background: In 2015, an international outbreak of Mycobacterium chimaera infections among patients undergoing cardiothoracic surgeries was associated with exposure to contaminated LivaNova 3T heater-cooler devices (HCDs). From June 2017 to October 2020, the Centers for Disease Control and Prevention was notified of 18 patients with M. chimaera infections who had undergone cardiothoracic surgeries at 2 hospitals in Kansas (14 patients) and California (4 patients); 17 had exposure to 3T HCDs. Whole-genome sequencing of the clinical and environmental isolates matched the global outbreak strain identified in 2015. Methods: Investigations were conducted at each hospital to determine the cause of ongoing infections. Investigative methods included query of microbiologic records to identify additional cases, medical chart review, observations of operating room setup, HCD use and maintenance practices, and collection of HCD and environmental samples. Results: Onsite observations identified deviations in the positioning and maintenance of the 3T HCDs from the US Food and Drug Administration (FDA) recommendations and the manufacturer’s updated cleaning and disinfection protocols. Additionally, most 3T HCDs had not undergone the recommended vacuum and sealing upgrades by the manufacturer to decrease the dispersal of M. chimaera–containing aerosols into the operating room, despite hospital requests to the manufacturer. Conclusions: These findings highlight the need for continued awareness of the risk of M. chimaera infections associated with 3T HCDs, even if the devices are newly manufactured. Hospitals should maintain vigilance in adhering to FDA recommendations and the manufacturer’s protocols and in identifying patients with potential M. chimaera infections with exposure to these devices.

Tracking the Distribution of Brucella abortus in Egypt Based on Core Genome SNP Analysis and In Silico MLVA-16

Brucellosis, caused by the bacteria of the genus Brucella, is one of the most neglected common zoonotic diseases globally with a public health significance and a high economic loss among the livestock industry worldwide. Since little is known about the distribution of B. abortus in Egypt, a total of 46 B. abortus isolates recovered between 2012–2020, plus one animal isolate from 2006, were analyzed by examining the whole core genome single nucleotide polymorphism (cgSNP) in comparison to the in silico multilocus variable number of tandem repeat analysis (MLVA). Both cgSNP analysis and MLVA revealed three clusters and one isolate only was distantly related to the others. One cluster identified a rather widely distributed outbreak strain which is repeatedly occurring for at least 16 years with marginal deviations in cgSNP analysis. The other cluster of isolates represents a rather newly introduced outbreak strain. A separate cluster comprised RB51 vaccine related strains, isolated from aborted material. The comparison with MLVA data sets from public databases reveals one near relative from Argentina to the oldest outbreak strain and a related strain from Spain to a newly introduced outbreak strain in Egypt. The distantly related isolate matches with a strain from Portugal in the MLVA profile. Based on cgSNP analysis the oldest outbreak strain clusters with strains from the UK. Compared to the in silico analysis of MLVA, cgSNP analysis using WGS data provides a much higher resolution of genotypes and, when correlated to the associated epidemiological metadata, cgSNP analysis allows the differentiation of outbreaks by defining different outbreak strains. In this respect, MLVA data are error-prone and can lead to incorrect interpretations of outbreak events.

Mycobacterium chimaera from Cardio Surgery Heating-Cooling Units and from Clinical Samples in Israel are Genetically Unrelated

Non-tuberculous mycobacteria (NTM) are opportunistic pathogens that cause illness primarily in the elderly, in the immunocompromised or in patients with underlying lung disease. Mycobacterium chimaera is a NTM species belonging to the M. avium Complex (MAC) group of species. Since 2013, a global outbreak of M. chimaera infection related to heater-cooler units (HCU) used in cardio-thoracic surgery has been identified. This outbreak was caused by a single strain of M. chimaera. In order to estimate the prevalence of this outbreak strain in Israel, we sampled M. chimaera from several HCU machines in Israel, as well as from patients, sequenced their genomes and compared them to the outbreak strain. The presence of mixed mycobacteria species in the samples complicated the analysis of obtained sequences. By applying a metagenomic binning strategy, we were able to obtain genomes of single strains from the mixed samples, and characterized them. M. chimaera strains were compared to each other and to previously reported genomes from other countries. The strain causing the outbreak related to the HCU machines was identified in several such machines in Israel but not in any of the clinical samples.

Genomic epidemiologic assessment implicates prolonged silent carriage, virulence factors and transmission between staff and patients in a NICU outbreak of MRSA.

Introduction: Methicillin-resistant Staphylococcus aureus (MRSA) is an important pathogen in neonatal intensive care units (NICU) that carries significant morbidity and mortality. Improving our understanding of MRSA transmission dynamics, especially among high risk patients, is an infection prevention priority. Methods: We investigated a cluster of clinical MRSA cases in the NICU using a combination of epidemiologic review and whole genome sequencing (WGS) of isolates from clinical and surveillance cultures obtained from patients and healthcare personnel (HCP). Results: Phylogenetic analysis identified two genetically distinct phylogenetic clades and revealed multiple silent transmission events between HCP and infants. The predominant outbreak strain harbored multiple virulence factors. Epidemiologic investigation and genomic analysis identified a HCP colonized with the dominant MRSA outbreak strain who cared for the majority of NICU patients who were infected or colonized with the same strain, including one NICU patient with severe infection seven months before the described outbreak. These results guided implementation of infection prevention interventions that prevented further transmission events. Conclusion: Silent transmission of MRSA between HCP and NICU patients likely contributed to a NICU outbreak involving a virulent MRSA strain. WGS enabled data-driven decision making to inform implementation of infection control policies that mitigated the outbreak. Prospective WGS coupled with epidemiologic analysis can be used to detect transmission events and prompt early implementation of control strategies.

Identification of the source of a Listeria monocytogenes outbreak by investigational tracing

The number of identified listeriosis outbreaks has increased since the sequence typing of Listeria monocytogenes isolates was established in Germany. Due to the nature of the disease, listeriosis outbreaks are difficult to solve. We present investigational tracing as a simple and rapid method to conduct outbreak investigations. The method was applied in 2019 to stop a prolonged listeriosis outbreak in Germany. The starting point for the investigational tracing was nine health care facilities (HCF). Single cases developed listeriosis while they were staying at the nine facilities. Data were collected from companies that delivered foods to HCF and from ready-to-eat (RTE) foods that were consumed there. Following a step-wise approach, data analysis identified similarities in the food supply of the HCF. Food data were heterogeneous and needed to be standardised. Own brands and changing article numbers were challenging aspects during the identification of manufacturers. The analysis of the delivering companies revealed no similarities. Detailed information about the consumed risk foods for Listeria contamination became available for six HCF. All facilities served a wide variety of cold cut meat products to their in-patients. Investigational tracing revealed that only meat products from one out of 29 food business operators had been consumed in all six HCF. Further activities of the authorities enabled the identification of the outbreak strain on food products and in the processing environment of this company. A product recall and the measures taken stopped the listeriosis outbreak. Thus, investigational tracing can be crucial for the clarification of listeriosis outbreaks.

A Series of Papaya-Associated Salmonella Illness Outbreak Investigations in 2017 and 2019 – A Focus on Traceback, Laboratory, and Collaborative Efforts

In 2017 and 2019, five outbreaks of infections from multiple strains of Salmonella linked to the consumption of whole, fresh Maradol papayas were reported in the United States, resulting in 325 ill persons. Traceback, laboratory, and epidemiologic evidence indicated papayas as the likely vehicle for each of these outbreaks and identified the source of papayas. State and FDA laboratories recovered Salmonella from papaya samples from various points of distribution, including at import entry, and conducted serotyping, pulsed-field gel electrophoresis (PFGE), and phylogenetic analyses of whole genome sequencing (WGS) data. Federal and state partners led traceback investigations to determine the source of papayas. Four different suppliers of papayas were linked by traceback and laboratory results to five separate outbreaks of Salmonella infections associated with papayas. In 2017, multiple states tested papaya samples collected at retail, and Maryland and Virginia investigators recovered strains of Salmonella associated with one outbreak. FDA collected 183 papaya samples in 2017, and 11 samples yielded 62 isolates of Salmonella. Eleven serotypes of Salmonella were recovered from FDA papaya samples, and nine serotypes were closely related genetically by PFGE and WGS to clinical isolates of four outbreaks, including the outbreak associated with positive state sample results. Four farms in Mexico were identified and their names were released to the general public, retailers, and foreign authorities. In 2019, FDA collected 119 papaya samples, three of which yielded Salmonella; none yielded the 2019 outbreak strain. Investigators determined that papayas of interest had been sourced from a single farm in Campeche, Mexico through traceback. This information was used to protect public health through public guidance, recalls, and import alerts and helped FDA collaborate with Mexican regulatory partners to enhance the food safety requirements for papayas imported from Mexico.

High frequency transmission, asymptomatic shedding, and airborne spread of Streptococcus pyogenes among schoolchildren exposed to scarlet fever: a longitudinal multi-cohort moleculo-epidemiological contact tracing study.

Background: Despite recommendations regarding prompt treatment of cases and enhanced hygiene measures, scarlet fever outbreaks increased in England between 2014-2018. Methods: We undertook a prospective, intensive contact tracing study in schools with consecutive scarlet fever cases to assess the impact of standard interventions on transmission of Streptococcus pyogenes between cases, classroom contacts, households, and classroom environments over 4 weeks using genome sequencing. Findings: Six classes, comprising 12 scarlet fever cases, 17 household contacts, and 278 classroom contacts were recruited. Prevalence of the outbreak strain in asymptomatic classroom contacts was high, increasing from 9.6% in week 1, to 26.9% in week 2, 23.9% in week 3, then 14.3% in week 4. Colonisation with non-outbreak strains was 0 - 7.5%. Genome sequencing showed clonality of isolates within each of six classes, confirming recent transmission accounted for high carriage. Of asymptomatic classroom contacts with S. pyogenes-positive throat swabs who were tested for transmissibility, 6/28 (21%) had positive cough plates and/or hand swabs, of whom three remained S. pyogenes-positive for 3 weeks. Only 1/60 surface swabs taken in 3 classrooms yielded S. pyogenes. In contrast, settle plates placed in elevated locations were S. pyogenes-positive in both classrooms tested. Interpretation: S. pyogenes transmission in schools is intense and may occur prior to, or in spite of reported treatment of cases, underlining a need for rapid case management. Despite guideline adherence, heavy shedding of S. pyogenes by small numbers of classroom contacts may perpetuate outbreaks, and airborne transmission has a plausible role in spread.