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ABSTRACT Salmonella enterica serovar Enteritidis is one of the most commonly reported serovars of nontyphoidal Salmonella causing human disease and is responsible for both gastroenteritis and invasive nontyphoidal Salmonella (iNTS) disease worldwide. Whole-genome sequence (WGS) comparison of Salmonella Enteritidis isolates from across the world has identified three distinct clades, global epidemic, Central/East African, and West African, all of which have been implicated in epidemics: the global epidemic clade was linked to poultry-associated gastroenteritis, while the two African clades were related to iNTS disease. However, the distribution and epidemiology of these clades across Africa are poorly understood because identification of these clades currently requires whole-genome sequencing capacity. Here, we report a sensitive, time- and cost-effective real-time PCR assay capable of differentiating between the Salmonella Enteritidis clades to facilitate surveillance and to inform public health responses. The assay described here is limited to previously confirmed S. Enteritidis isolates. IMPORTANCE Challenges in the diagnosis and treatment of invasive Salmonella Enteritidis bloodstream infections in sub-Saharan Africa are responsible for a case fatality rate of approximately 15%. It is important to identify distinct clades of S. Enteritidis in diagnostic laboratories in the African setting to determine the different health outcomes associated with particular outbreaks. Here, we describe the development of a high-quality molecular classification assay for clade typing of S. Enteritidis that is ideal for use in public health laboratories in resource-limited settings.

INTRODUCTION: In 2009 and 2010, more than 6,000 cholera cases were recorded during these outbreaks with more than 80% of cases recorded in Lusaka province. After a five-year break, in 2016 an outbreak occurred in Lusaka, causing more than 1,000 cases of cholera. This study will strengthen the epidemiological information on the changing characteristics of the cholera outbreaks, for treatment, prevention and control of the disease. METHODS: this was a laboratory-based descriptive cross-sectional study conducted at the University Teaching Hospital in Lusaka, Zambia. A total of 83 V. cholerae O1 isolates were characterised by biochemical testing, serotyping, antimicrobial susceptibility testing, and macrorestriction analysis using Pulsed-Field Gel Electrophoresis. RESULTS: macrorestriction analysis of the isolates demonstrated high genetic diversity among the isolates with 16 different patterns. The largest pattern comprised 9 isolates while the smallest one had 1 isolate. 2009 and 2010 isolates were highly resistant to nalidixic acid and cotrimoxazole, but highly sensitive to azithromycin and ampicillin. Of the fifty-two isolates from the 2016 cholera outbreak, 90% (47) were sensitive to cotrimoxazole, 94% (49) to tetracycline, and 98% (51) to azithromycin, while 98% (51) were resistant to nalidixic acid and 31(60%) to ampicillin. CONCLUSION: macrorestriction analysis demonstrated high genetic diversity among the V. cholerae O1 strains, suggesting that these isolates were probably not from a similar source. This study also revealed the emergence of multidrug resistance among the 2016 V. cholerae outbreak isolates but were susceptible to cotrimoxazole, tetracycline, and azithromycin, which can be used for treatment of the cholera cases.

Abstract Background Salmonella enterica serovar Enteritidis (Salmonella Enteritidis) has become a significant pathogen in South Africa, and the need for improved molecular surveillance of this pathogen has become important. Over the years, multi-locus variable-number tandem-repeats analysis (MLVA) has become a valuable molecular subtyping technique for Salmonella, particularly for highly homogenic serotypes such as Salmonella Enteritidis. This study describes the use of MLVA in the molecular epidemiological investigation of outbreak isolates in South Africa. Methods Between the years 2013 and 2015, the Centre for Enteric Diseases (CED) received 39 Salmonella Enteritidis isolates from seven foodborne illness outbreaks, which occurred in six provinces. MLVA was performed on all isolates. Results Three MLVA profiles (MLVA profiles 21, 22 and 28) were identified among the 39 isolates. MLVA profile 28 accounted for 77% (30/39) of the isolates. Isolates from a single outbreak were grouped into a single MLVA profile. A minimum spanning tree (MST) created from the MLVA data showed a close relationship between MLVA profiles 21, 22 and 28, with a single VNTR locus difference between them. Conclusions MLVA has proven to be a reliable method for the molecular epidemiological investigation of Salmonella Enteritidis outbreaks in South Africa. These foodborne outbreaks emphasize the importance of the One Health approach as an essential component for combating the spread of zoonotic pathogens such as Salmonella Enteritidis.

Abstract Background Workers in clinical microbiology laboratories are exposed to a variety of pathogenic microorganisms. Salmonella species is among the most commonly reported bacterial causes of laboratory-acquired infections. We report on three cases of laboratory-acquired Salmonella enterica serotype Typhi (Salmonella Typhi) infection which occurred over the period 2012 to 2016 in South Africa. Methods Laboratory investigation included phenotypic and genotypic characterization of isolates. Phenotypic analysis included standard microbiological identification techniques, serotyping and antimicrobial susceptibility testing. Genotypic analysis included the molecular subtyping methodologies of pulsed-field gel electrophoresis analysis, multilocus sequence typing and whole-genome sequencing (WGS); with WGS data analysis including phylogenetic analysis based upon comparison of single nucleotide polymorphism profiles of isolates. Results All cases of laboratory-acquired infection were most likely the result of lapses in good laboratory practice and laboratory safety. The following critical issues were highlighted. There was misdiagnosis and misreporting of Salmonella Typhi as nontyphoidal Salmonella by a diagnostic laboratory, with associated public health implications. We highlight issues concerning the importance of accurate fluoroquinolone susceptibility testing and interpretation of results according to updated guidelines. We describe potential shortcomings of a single disk susceptibility screening test for fluoroquinolone susceptibility and suggest that confirmatory minimum inhibitory concentration testing should always be performed in cases of invasive Salmonella infections. These antimicrobial susceptibility testing issues resulted in inappropriate ciprofloxacin therapy which may have been responsible for failure in clearance of pathogen from patients. Salmonella Typhi capsular polysaccharide vaccine was not protective in one case, possibly secondarily to a faulty vaccine. Conclusions Molecular subtyping of isolates proved effective to investigate the genetic relatedness of isolates. Molecular subtyping data interpreted together with epidemiological data allowed us to pinpoint the most likely sources for our cases of laboratory-acquired infection.

BACKGROUND: Drug resistance of Salmonella enterica serovar Typhi (Salmonella Typhi) to first-line antibiotics is emerging in Central Africa. Although increased use of fluoroquinolones is associated with spread of resistance, Salmonella Typhi with decreased ciprofloxacin susceptibility (DCS) has rarely been reported in Central Africa. METHODOLOGY/PRINCIPAL FINDINGS: As part of a microbiological surveillance study in the Democratic Republic of the Congo (DR Congo), Salmonella Typhi isolates from bloodstream infections were collected prospectively between 2007 and 2011. The genetic relationship of the Salmonella Typhi isolates was assessed by pulsed-field gel electrophoresis (PFGE). The antimicrobial resistance profile of the isolates was determined and mutations associated with DCS were studied. In total, 201 Salmonella Typhi isolates were collected. More than half of the Salmonella Typhi isolates originated from children and young adults aged 5-19. Thirty different PFGE profiles were identified, with 72% of the isolates showing a single profile. Multidrug resistance, DCS and azithromycin resistance were 30.3%, 15.4% and 1.0%, respectively. DCS was associated with point mutations in the gyrA gene at codons 83 and 87. CONCLUSIONS/SIGNIFICANCE: Our study describes the first report of widespread multidrug resistance and DCS among Salmonella Typhi isolates from DR Congo. Our findings highlight the need for increased microbiological diagnosis and surveillance in DR Congo, being a prerequisite for rational use of antimicrobials and the development of standard treatment guidelines.