Impact of intense sanitization on environmental biofilm communities and the survival of Salmonella enterica at a beef processing plant.

Salmonella enterica is a leading cause of foodborne illness in the U.S. In the meat industry, one action taken to address pathogen contamination incidence is an intense sanitization (IS) of the entire processing plant that many large processors perform annually or semiannually. However, this procedure's immediate and long-term impact on environment microbial community and pathogen colonization are unknown. Here we investigated the impact of IS procedure on environmental biofilms and the subsequent S. enterica colonization and stress tolerance. Environmental samples were collected from floor drains at various areas 1 week before, 1 week, and 4 weeks after the IS procedure at a beef plant with sporadic S. enterica prevalence. Biofilm formation by microorganisms in the drain samples without S. enterica presence was tested under processing temperature. The ability of the biofilms to recruit and/or protect a co-inoculated S. enterica strain from quaternary ammonium compound (QAC) treatment was determined. The community structure of each drain sample was elucidated through 16S rRNA amplicon community sequencing. Post-IS samples collected from 8 drains formed significantly stronger biofilms than the respective pre-IS samples. S. enterica colonization was not different between the pre- and post-IS biofilms at all drain locations. S. enterica survival in QAC-treated pre- and post-IS mixed biofilms varied depending upon the drain location but a higher survival was associated with a stronger biofilm matrix. The 16S rRNA amplicon gene community sequencing results exhibited a decrease in community diversity 1 week after IS treatment but followed by a significant increase 4 weeks after the treatment. The IS procedure also significantly altered the community composition and the higher presence of certain species in the post-IS community may be associated with the stronger mixed biofilm formation and Salmonella tolerance. Our study suggested that the IS procedure might disrupt the existing environmental microbial community and alter the natural population composition, which might lead to unintended consequences as a result of a lack of competition within the multispecies mixture. The survival and recruitment of species with high colonizing capability to the post-IS community may play crucial roles in shaping the ensuing ecological dynamics.

Salmonella-induced microbiome profile in response to sanitation by quaternary ammonium chloride.

Salmonella enterica is a prominent cause of foodborne disease in the United States. However, the mechanism and route of pathogen transmission that leads to Salmonella infection in commercial processing plants are poorly understood. This study aimed to investigate the effect of mixed-species biofilms on S. enterica survival and persistence under sanitizer stress [Quaternary ammonium compounds (QACs)] by analyzing 78 floor drain samples from a meat processing facility and three S. enterica strains (serovars Cerro, Montevideo, and Typhimurium) isolated from that facility and an unrelated source. The four test groups were as follows: control, QAC treatment, Salmonella addition, and QAC treatment with Salmonella addition. DNAs were extracted, and 16S rRNA gene based on the variable region V4 amplicon sequencing was performed to analyze the relative abundance, core microbiome, and Alpha and Beta diversity using the qiime2 pipeline. At the genus level, the Brochothrix (45.56%), Pseudomonas (38.94%), Carnobacterium (6.18%), Lactococcus (4.68%), Serratia (3.14%), and Staphylococcus (0.82%) were shown to be the most prevalent in all drain samples. The results demonstrate that the relative abundance of different bacterial genera was affected by both QAC treatment and Salmonella addition, with some genera showing increases or decreases in abundance. Notably, the correlation network was constructed to understand the relationships between the different bacteria. Nitrospira had the greatest number of connections in the floor drain environment network, with two negative and eight positive correlations. The results suggest that Nitrospira in the mixed-species biofilm community may play a role in converting ammonium in the QAC sanitizer into nitrites. Thus, Nitrospira could be a potentially important genus in providing sanitizer resistance to pathogen-encompassed mixed-species biofilms.IMPORTANCESalmonella contamination in meat processing facilities can lead to foodborne illness outbreaks. Our study characterized the microbiome dynamics in beef facility drains and their response to Salmonella addition and common sanitizer (QAC). Nitrospira could be an important genus in providing sanitizer resistance to pathogen-encompassed mixed-species biofilms. The results provide insight into the impact of mixed-species biofilms on Salmonella survival and persistence under sanitizer stress in meat processing facilities. The results highlight the need to consider mixed-species biofilm effects when developing targeted interventions to enhance food safety.

Shotgun-metagenomics reveals a highly diverse and communal microbial network present in the drains of three beef-processing plants.

Background: Multi-species biofilms pose a problem in various environments, especially food-processing environments. The diversity of microorganisms in these biofilms plays a critical role in their integrity and protection against external biotic and abiotic factors. Compared to single-species biofilms, mixed-species biofilms are more resistant to various stresses, including antimicrobials like sanitizers. Therefore, understanding the microbiome composition and diversity in biofilms and their metabolic potential is a priority when developing intervention techniques to combat foodborne pathogens in food processing environments. Methods: This study aimed to describe and compare the microbiome profile of 75 drain biofilm samples obtained from five different locations (Hotscale, Hotbox, Cooler, Processing, & Grind room) of three beef-processing plants (Plant A, B & C) taken over two timepoints 2017-18 (T1) and 2021 (T2) by shotgun sequencing. Results: Core microbiome analysis found Pseudomonas, Psychrobacter, and Acinetobacter to be the top three prevalent genera among the plants and locations. Alpha diversity analysis demonstrated a high diversity of microbiome present in all the plants and locations across the time points. Functional analysis showed the high metabolic potential of the microbial community with abundance of genes in metabolism, cell-adhesion, motility, and quorum sensing. Moreover, Quaternary Ammonium Compound (QAC) resistance genes were also observed, this is significant as QAC sanitizers are commonly used in many food processing facilities. Multi-functional genes such as transposases, polymerases, permeases, flagellar proteins, and Mobile Genetic Elements (MGEs) were found suggesting these are dynamic microbial communities that work together to protect themselves against environmental stresses through multiple defense mechanisms. Conclusion: This study provides a framework for understanding the collective microbial network spanning a beef processing system. The results can be used to develop intervention strategies to best control these highly communicative microbial networks.

Characterization of boar semen microbiome and association with sperm quality parameters.

Elevated levels of bacteria within fresh extended boar semen are associated with decreased sperm longevity, therefore reducing the fertility of a semen dose. The objective of this study was to characterize the bacterial communities using 16S rRNA sequencing in freshly extended boar semen samples and relate the prevalence and diversity of the microbial population to sperm quality parameters 1) between studs, 2) between pooled and single-sire doses, and 3) over a 5-day period. Eight single-sire (n = 4 per stud) and eight pooled (n = 4 per stud) non-frozen extended semen doses were obtained from two boar studs (A and B). Pooled doses were the composite of the boar's ejaculates used in single-sire doses. Doses were subsampled for 5 d post-collection. Ten negative controls of each pooled dose (n = 2) and single-sire dose (n = 8) remained sealed until the last day. Microbiome analysis was achieved by examining the V4 hypervariable region of the 16S rRNA gene of flash-frozen samples. Two evaluators determined the average sperm motility and agglutination (0: no adhesion to 3: >50% adhesion) by averaging their estimates together at 10 random locations per slide. Stud A had greater sperm agglutination (1.6 vs. 1.0 ± 0.1; P < 0.01) than stud B. Sperm motility decreased over the 5-day period (P < 0.01) and tended (P = 0.09) to be greater in stud B than A (67.4% vs. 61.5% ± 0.02%). Compared with stud A, stud B had a greater relative abundance of Proteobacteria (60.0% vs. 47.2% ± 1.5%; P < 0.01) and a lower relative abundance of Firmicutes (22.5% vs. 31.9% ± 1.4%; P < 0.01). Moreover, stud A had a greater relative abundance of Bacteroidetes (6.3% vs. 5.3% ± 0.4%; P < 0.01) and Actinobacteria (11.5% vs. 10.1% ± 0.5%; P = 0.05) than stud B. Differences were found in alpha diversity for both Chao1 (P < 0.01) and Shannon (P < 0.01) diversity indexes among days 2, 3, 4, and 5 post-collection to day 1. For beta diversity, unweighted UniFrac metric on days 2, 3, 4, and 5 post-collection differed from those on day 1 (P < 0.01). There were significant correlations between sperm motility and relative abundance of Prevotella (r = -0.29), Ruminococcus (r = -0.24), and Bacteroides (r = -0.32). Additionally, there were significant correlations between sperm motility and Chao1 (r = -0.50) and Shannon's index (r = -0.36). These results demonstrate that differences in bacterial communities over time and between boar studs can be associated with variation in sperm quality.

The ability to improve production output remains essential to meet the growing global demand for pork. However, the presence of pathogenic bacteria such as Escherichia coli and Clostridium perfringens can hinder production goals by reducing semen quality through increased clumping events and decreased sperm motility. In addition, reduced conception rates and decreased litter size can occur when bacterial-contaminated semen doses are used for artificial insemination. The purpose of this study was to determine the bacterial communities within freshly extended boar semen and associate specific bacterial communities with sperm quality measurements. Current findings suggest that certain bacteria can accumulate within a group of animals or over a short period of time which can impact sperm cell characteristics. Having less diverse bacterial communities also appears to be associated with favorable semen quality. Future research is needed to determine the interactions different bacterial communities have with sperm cells to characterize their nature as pathogenic or commensal.

Prevalence of differential microbiome in healthy, diseased and nipped colonies of corals, Porites lutea in the Gulf of Kachchh, north-west coast of India.

Coral reefs are constantly subjected to multiple stresses like diseases and fish predation, which can profoundly influence the coral microbiome. This study investigated the differences in bacterial community structure of healthy, white syndrome affected and blenny nipped coral colonies of Porites lutea, collected from the coral reefs of Gulf of Kachchh, north-west coast of India. Present study observed that the stressed coral colonies harbored more OTUs and contained higher diversity values compared to healthy corals colonies. Similarly, beta diversity analysis indicated the dissimilarities among the three coral samples analyzed. Though the taxonomy analysis indicated bacterial phyla like Proteobacteria, Firmicutes, Bacteroidetes, and Actinobacteria among the entire coral samples studied, there was a variation in their relative abundances. Huge variations were observed in the relative dominance at the bacterial genera level. About 13phyla and 11 genera was identified in healthy coral. The PBN sample was found to contain Proteobacteria, Cyanobacteria, Verrucomicrobia, and Lentisphaerae as dominant phyla and Endozoicomonas, Dyella, Woeseia, and Winogradskyella as dominant genera. The PWS sample contained Proteobacteria, Lentisphaerae, Spirochaetes, and Tenericutes as dominant phyla and Endozoicomonas, Arcobacter, Sunxiuqinia, and Carboxylicivirgia as dominant genera. Among the healthy samples, sequences belonging to Uncultured Rhodospirillaceae were dominant, while Woeseia and sequences belonging to Uncultured Rhodovibrionaceae were dominant among the blenny nipped white syndrome infected corals. Although any previously established pathogen was not identified, present study revealed the presence of a potentially pathogenic bacterium, Arcobacter, among the diseased corals. It also demonstrated a dynamic microbiome among the Porites lutea colonies on subjecting to various stresses.

Uncommon cause of acute appendiceal torsion mucocele.

Appendiceal torsion is a rare phenomenon, presenting with symptoms mimicking acute appendicitis. Preoperative imaging generally has a low pick-up rate of appendicular torsion and is thus mostly diagnosed intraoperatively. The cause of the appendicular torsion should guide the required surgical resection to ensure adequate oncological treatment. Therefore, emphasis should be placed on minimal tissue handling in such operative cases to reduce the risk of accidental rupture, tumour spillage and spread.

Effect of chlorpyrifos on the earthworm Eudrilus euginae and their gut microbiome by toxicological and metagenomic analysis.

The earthworms are important soil invertebrates and play a crucial role in pedogenesis. The application of pesticides and prolonged exposure to pesticides causes mortality of earthworms apart from profoundly affecting the resident gut microbiome. The microbiome plays a significant effect on the metabolic processes associated with earthworms. The pesticide Chlorpyrifos (CPF) was studied for its toxicity on Eudrilus euginae by toxicity studies. The LC50 value of filter paper contact test and acute toxicity test was 3.8 mg/mL and 180 mg/kg. The prolonged exposure of earthworms to pesticide on reproductive toxicity resulted in the mortality of earthworms and absence of cocoon formation. Further, the effects of CPF on the whole gut microbiome of E. euginae was analyzed using a long amplicon Nanopore sequencing. Results indicated no fluctuations with Firmicutes and Bacteroidetes, that were found to be dominant at bacterial phyla level while at the genus level, remarkable differences were noticed. Clostridium dominated the earthworm gut prior to CPF exposure while Bacillus dominated after exposure. Similarly, the fungal members such as Ascomycota and Basidiomycota were observed to dominate the gut of earthworm at the phyla level before and after exposure to CPF. In contrast, Clavispora (65%) was the dominant genus before CPF exposure and Taloromyces (42%) dominated after the CPF exposure. Our study demonstrates the effect of CPF on the mortality of E. euginae while the amplicon sequencing established the unique microbiome of the gut in response to the CPF exposure.

Correlation of dental age, skeletal age, and chronological age among children aged 9-14 years: A retrospective study.

BACKGROUND: Growth can be one of the most uncertain variations, but understanding the same is very important for diagnosis and treatment planning. Skeletal age and dental age have been used to determine a child's developmental age. Several researchers have evaluated the association between dental and skeletal maturity with chronologic age on different population. The purpose of the present study was to find out whether dental age estimation can be replaced for skeletal age estimation in the Dakshina Kannada population. METHODS: A total of 104 samples equally distributed among both genders in the age group of 9-14 years were selected. Skeletal age was estimated using hand-wrist radiographs and Fishman's skeletal maturation index and dental age was measured using Demirjian's method. RESULTS: Spearman's rank-order correlation coefficients were used to measure the association between the skeletal maturity and dental maturity. The mean ages of male and female samples were determined as 11.89 ± 1.867 years and 12.21 ± 1.473 years, respectively. Chronological age was found to be positively correlated to dental age and skeletal age and found to be statistically significant (P < 0.01). The correlation between dental age and skeletal age was also found to be statistically significant with P < 0.001 and correlation coefficient of 0.683 and 0.704 for males and females. CONCLUSION: The present study showed a strong relation between the developmental ages in mixed dentition population; hence, dental age can be considered as a replacement in the study population.