Toward the Adoption of Loop-Mediated Isothermal Amplification for Salmonella Screening at the National Antimicrobial Resistance Monitoring System's Retail Meat Sites.

The National Antimicrobial Resistance Monitoring System (NARMS) is a One Health program in the United States that collects data on antimicrobial resistance in enteric bacteria from humans, animals, and the environment. Salmonella is a major pathogen tracked by the NARMS retail meat arm but currently lacks a uniform screening method. We evaluated a loop-mediated isothermal amplification (LAMP) assay for the rapid screening of Salmonella from 69 NARMS retail meat and poultry samples. All samples were processed side by side for culture isolation using two protocols, one from NARMS and the other one described in the U.S. Food and Drug Administration's Bacteriological Analytical Manual (BAM). Overall, 10 (14.5%) samples screened positive by the Salmonella LAMP assay. Of those, six were culture-confirmed by the NARMS protocol and six by the BAM method with overlap on four samples. No Salmonella isolates were recovered from samples that screened negative with LAMP. These results suggested 100% sensitivity for LAMP in reference to culture. Antimicrobial susceptibility testing and whole-genome sequencing analysis confirmed identities of these isolates. Using the BAM protocol, all Salmonella isolates were recovered from samples undergoing Rappaport-Vassiliadis medium selective enrichment and presumptive colonies (n = 130) were dominated by Hafnia alvei (44.6%), Proteus mirabilis (22.3%), and Morganella morganii (9.9%) based on matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. This method comparison study clearly demonstrated the benefit of a rapid, robust, and highly sensitive molecular screening method in streamlining the laboratory workflow. Fourteen NARMS retail meat sites further verified the performance of this assay using a portion of their routine samples, reporting an overall specificity of 98.8% and sensitivity of 90%. As of July 2022, the vast majority of NARMS retail meat sites have adopted the Salmonella LAMP assay for rapid screening of Salmonella in all samples.

Surgical transversus abdominis plane block with liposomal bupivacaine at cesarean: a pilot randomized trial.

BACKGROUND: Postcesarean pain control is challenging. In addition to intrathecal morphine, recent studies have shown that liposomal bupivacaine administered via conventional transversus abdominis plane block reduces postcesarean opioid use. However, whether the administration of liposomal bupivacaine via a surgical approach also reduces opioid use is unknown. OBJECTIVE: This study aimed to investigate whether the administration of liposomal bupivacaine via surgical transversus abdominis plane block (TAP block) reduces the cumulative dose of opioids administered in the first 48 hours after cesarean delivery among participants who also receive intrathecal morphine. STUDY DESIGN: This was a pilot single-blind randomized controlled trial of 60 parturients undergoing cesarean delivery at a community tertiary referral hospital staffed by academic physicians. Immediately before fascial closure during cesarean delivery, a total of 80 mL of dilute bupivacaine plus liposomal bupivacaine or dilute bupivacaine alone was administered via surgical transversus abdominis plane block (40 mL on each side). The primary outcome was a median cumulative opioid dose received within the first 48 hours after cesarean delivery measured in morphine milligram equivalents. In addition, opioid use at other time points, pain scores, and participant satisfaction were assessed. A sample size of 60 was determined to be adequate to inform a potential future adequately powered randomized trial. The primary outcome of morphine milligram equivalents and pain scores were compared using a Wilcoxon rank-sum test. RESULTS: Between October 11, 2021, and August 29, 2022, 60 participants were randomized and analyzed: 31 were allocated to liposomal bupivacaine plus regular bupivacaine (intervention group), and 29 were allocated to regular bupivacaine alone (control group). Participants allocated to the intervention group used a median cumulative dose of 2 morphine milligram equivalents of opioids (interquartile range, 0-24) in the first 48 hours compared with 8 morphine milligram equivalents (interquartile range, 0-40) among participants allocated to the control group (P=.236). The percentage of participants who used ≤15 morphine milligram equivalents of opioids was 61% in the intervention arm and 41% in the control arm (P=.123), and the percentage who used zero opioids was 45% in the intervention arm and 34% in the control arm (P=.399). The total number of opioid pills prescribed at discharge was fewer in the intervention arm than in the control arm (P=.029). Patient satisfaction with the intervention group and control group was similar. CONCLUSION: Our pilot study suggests that liposomal bupivacaine administered via surgical transversus abdominis plane block is worth critical evaluation as an adjunctive analgesic modality in an adequately powered randomized trial.

Versatility of a Salmonella Loop-Mediated Isothermal Amplification Assay Using Multiple Platforms and Master Mixes in Animal Food Matrices.

BACKGROUND: Improvement in Salmonella detection methods greatly enhances the efficiency of various food testing programs. A Salmonella loop-mediated isothermal amplification (LAMP) assay has been validated in animal food through multi-laboratory validation. OBJECTIVE: The study aimed to demonstrate the versatility of this molecular assay while expanding it to multiple platforms and various reagent choices for use in animal food testing. METHODS: Following the U.S. Food and Drug Administration (FDA)'s Guidelines for the Validation of Analytical Methods for the Detection of Microbial Pathogens in Foods and Feeds, we examined the inclusivity, exclusivity, and LOD of the assay using two platforms (7500 Fast and Genie II) and three LAMP master mixes (GspSSD, GspSSD2.0, and WarmStart) in seven animal food matrixes (dry cat food, dry dog food, cattle feed, dairy feed, horse feed, poultry feed, and swine feed). The FDA's Bacteriological Analytical Manual (BAM) Salmonella culture method was the reference method. RESULTS: Inclusivity and exclusivity data were consistent among all six platform and master mix combinations with a few exceptions. Comparable LODs were observed down to the single-cell level (WarmStart was 10-fold less sensitive). Performance was similar to the BAM method for detecting fractional positive results in seven animal food matrixes. Nonetheless, LAMP time to positive results and annealing/melting temperature differed among master mixes and platforms. CONCLUSION: The Salmonella LAMP assay was successfully validated in two platforms and three master mixes, making it a flexible tool for use by the FDA's field laboratories in regulatory testing of animal food and for adoption by other food testing programs. HIGHLIGHTS: We demonstrated the LAMP assay's versatility on two platforms and three master mixes for the rapid and reliable screening of Salmonella in seven animal food matrixes. GspSSD2.0 was the fastest master mix (time to positive results as early as 3.5 min) while Genie II had several attractive features from a user perspective.

Chromosome-level genome assembly of the blue crab, Callinectes sapidus.

The blue crab, Callinectes sapidus (Rathbun, 1896) is an economically, culturally, and ecologically important species found across the temperate and tropical North and South American Atlantic coast. A reference genome will enable research for this high-value species. Initial assembly combined 200× coverage Illumina paired-end reads, a 60× 8 kb mate-paired library, and 50× PacBio data using the MaSuRCA assembler resulting in a 985 Mb assembly with a scaffold N50 of 153 kb. Dovetail Chicago and HiC sequencing with the 3d DNA assembler and Juicebox assembly tools were then used for chromosome scaffolding. The 50 largest scaffolds span 810 Mb are 1.5-37 Mb long and have a repeat content of 36%. The 190 Mb unplaced sequence is in 3921 sequences over 10 kb with a repeat content of 68%. The final assembly N50 is 18.9 Mb for scaffolds and 9317 bases for contigs. Of arthropod BUSCO, ∼88% (888/1013) were complete and single copies. Using 309 million RNAseq read pairs from 12 different tissues and developmental stages, 25,249 protein-coding genes were predicted. Between C. sapidus and Portunus trituberculatus genomes, 41 of 50 large scaffolds had high nucleotide identity and protein-coding synteny, but 9 scaffolds in both assemblies were not clear matches. The protein-coding genes included 9423 one-to-one putative orthologs, of which 7165 were syntenic between the two crab species. Overall, the two crab genome assemblies show strong similarities at the nucleotide, protein, and chromosome level and verify the blue crab genome as an excellent reference for this important seafood species.

Corrigendum: Effect of Diet on the Enteric Microbiome of the Wood-Eating Catfish Panaque nigrolineatus.

[This corrects the article DOI: 10.3389/fmicb.2019.02687.].

Effect of Diet on the Enteric Microbiome of the Wood-Eating Catfish Panaque nigrolineatus.

Wood is consistently found in high levels in the gastrointestinal tract of the Amazonian catfish Panaque nigrolineatus, which, depending on environmental conditions, can switch between xylivorous and detritivorous dietary strategies. This is highly unusual among primary wood consumers and provides a unique system to examine the effect of dietary change in a xylivorous system. In this study, microbiome and predictive metagenomic analyses were performed for P. nigrolineatus fed either wood alone or a less refractory mixed diet containing wood and plant nutrition. While diet had an impact on enteric bacterial community composition, there was a high degree of interindividual variability. Members of the Proteobacteria and Planctomycetes were ubiquitous and dominated most communities; Bacteroidetes, Fusobacteria, Actinobacteria, and Verrucomicrobia also contributed in a tissue and diet-specific manner. Although predictive metagenomics revealed functional differences between communities, the relative abundance of predicted lignocellulose-active enzymes remained similar across diets. The microbiomes from both diets appeared highly adapted for hemicellulose hydrolysis as the predicted metagenomes contained several classes of hemicellulases and lignin-modifying enzymes. Enteric communities from both diets appeared to lack the necessary cellobiohydrolases for efficient cellulose hydrolysis, suggesting that cellobiose is not the primary source of dietary carbon for the fish. Our findings suggest that the P. nigrolineatus gut environment selects for an enteric community based on function, rather than a vertically transferred symbiotic relationship. This functional selection strategy may provide an advantage to an organism that switches between dietary strategies to survive a highly variable environment.