Natural products biosynthesized wholly or in part by nonribosomal peptide synthetases (NRPSs) are some of the most important drugs currently used clinically for the treatment of a variety of diseases. Since the initial research into NRPSs in the early 1960s, we have gained considerable insights into the mechanism by which these enzymes assemble these natural products. This review will present a brief history of how the basic mechanistic steps of NRPSs were initially deciphered and how this information has led us to understand how nature modified these systems to generate the enormous structural diversity seen in nonribosomal peptides. This review will also briefly discuss how drug development and discovery are being influenced by what we have learned from nature about nonribosomal peptide biosynthesis.

Several Cronobacter species are opportunistic pathogens that cause infections in humans. The aim of this study was to detect Cronobacter spp. from 90 samples of retail foods in Brazil, and characterize the strains by phenotypic tests, molecular assays and antibiotic susceptibility. Three isolation methodologies were evaluated using different selective enrichments and the isolates were identified using Vitek 2.0, PCRs protocols, fusA allele sequencing and multilocus sequence typing (MLST). Thirty-eight samples (42.2%) contained Cronobacter spp., and the highest percentage was found in flours (66.7%, 20/30), followed by spices and herbs (36.7%, 11/30), and cereal mixes for children (23.3%, 7/30). The 45 isolates included four species: C. sakazakii (n = 37), C. malonaticus (n = 3), C. dublinensis (n = 3), and C. muytjensii (n = 2); that presented 20 different fusA alleles. MLST analysis revealed 32 sequence types (STs), 13 of which were newly identified. All strains were sensitive to all antibiotics (n = 10) tested. The combination of CSB/v enrichment with DFI plating was considered the most efficient for Cronobacter spp. isolation. This study revealed the presence of Cronobacter spp. in foods commercialized in Brazil and the isolates showed a high diversity after MLST analysis and included two strains of the C. sakazakii ST4 neonatal meningitic pathovar.

In 2010, two infants became ill at a hospital in Mexico. Subsequently, a range of clinical, environmental, and powdered and rehydrated infant formula isolates were identified by using a combination of phenotyping and PCR probes. The strains were clustered according to pulsed-field gel electrophoresis. The causative agent was reported as Cronobacter sakazakii, with powdered infant formula (PIF) identified as the likely source of the infections. This new study further characterized the isolates from this outbreak by using multilocus sequence typing and whole genome sequencing of selected strains. Though four PIF isolates and one hospital environmental isolate were identified as C. sakazakii sequence type 297 by multilocus sequence typing, they were isolated 6 months prior to the outbreak. Genotypic analyses of patient isolates identified them as Enterobacter hormaechei and Enterobacter spp. The pulsed-field gel electrophoresis profile of the Enterobacter spp. isolates matched those of isolates from previously unopened tins of PIF. E. hormaechei was only isolated from the two infants and not PIF. The reevaluation of this outbreak highlights the need for accurate detection and identification assays, particularly during outbreak investigations in which incorrect identifications may mislead the investigation and attribution of the source. Though the species responsible for the symptoms could not be determined, this outbreak demonstrated the possible transmission of Enterobacter spp. from PIF to infants. These are possibly the first reported cases of Enterobacter spp. infection of infants from bacterial-contaminated PIF.

A nationwide survey was conducted to obtain qualitative and quantitative data on bacterial contamination of raw commingled silo milk intended for pasteurization. The levels of total aerobic bacteria, total coliforms, Enterobacteriaceae, Escherichia coli, and Staphylococcus aureus were determined using the TEMPO system. The prevalence rates and levels of presumptive Bacillus cereus, E. coli O157:H7, Listeria monocytogenes, and Salmonella spp. were determined in 214 samples. B. cereus was detected in 8.91% of samples, at 3.0 to 93 CFU/ml. E. coli O157:H7 was detected in 3.79 to 9.05% of samples, at <0.0055 to 1.1 CFU/ml, depending on the assay utilized. Salmonella spp. were recovered from 21.96 to 57.94% of samples, at <0.0055 to 60 CFU/ml. L. monocytogenes was detected in 50.00% of samples, at <0.0055 to 30 CFU/ml. The average log-transformed counts of total viable bacteria were slightly lower in samples containing no pathogens. No correlation was observed between the levels of organisms detected with the TEMPO system and the presence or levels of any pathogen except E. coli O157:H7. A higher average log-transformed count of total viable bacteria was observed in samples positive for this organism. The high prevalence rates of target pathogens may be attributed to a variety of factors, including detection methods, sample size, and commingling of the milk in the silo. The effects of commingling likely contributed to the high prevalence rates and low levels of target pathogens because of the inclusion of milk from multiple bulk tanks. The high prevalence rates also may be the result of analysis of larger sample volumes using more sensitive detection methods. These quantitative data could be utilized to perform more accurate risk assessments and to better estimate the appropriate level of protection for dairy products and processing technologies.