Maintenance of the Shigella sonnei virulence plasmid is dependent on its repertoire and amino acid sequence of toxin:antitoxin systems

Shigella sonnei is a major cause of bacillary dysentery, and of increasing concern due to the spread of multi-drug resistance. S. sonnei harbours pINV, a ∼ 210 kb plasmid that encodes a Type III secretion system (T3SS), which is essential for virulence. During growth in the laboratory, avirulence arises spontaneously in S. sonnei at high frequency, hampering studies on and vaccine development against this important pathogen. Here we investigated the molecular basis for the emergence of avirulence in S. sonnei , and show that avirulence mainly results from pINV loss, consistent with previous findings. Ancestral deletions have led to the loss from S. sonnei pINV of two toxin:antitoxin (TA) systems involved in plasmid maintenance, CcdAB and GmvAT, which are found on pINV in Shigella flexneri . We show that introduction of these TA systems into S. sonnei pINV reduced but did not eliminate pINV loss, while single amino acid polymorphisms found in the S. sonnei VapBC TA system compared with S. flexneri VapBC also contribute to pINV loss. Avirulence also results from deletions of T3SS-associated genes on pINV through recombination between insertion sequences (ISs) on the plasmid; these events differ from those observed in S. flexneri due to the different distribution and repertoire of ISs. Our findings demonstrate that TA systems and ISs influence plasmid dynamics and loss in S. sonnei , and could be exploited for the design and evaluation of vaccines. IMPORTANCE Shigella sonnei is the major cause of shigellosis in high-income and industrialising countries, and an emerging multi-drug resistant pathogen. A significant challenge when studying this bacterium is that it spontaneously becomes avirulent during growth in the laboratory, through loss of its virulence plasmid (pINV). Here we decipher the mechanisms leading to avirulence in S. sonnei and how the limited repertoire and amino acid sequences of plasmid-encoded toxin:antitoxin (TA) systems make the maintenance of pINV in this bacterium less efficient compared with Shigella flexneri . Our findings highlight how subtle differences in plasmids in closely-related species have marked effects and could be exploited to reduce plasmid loss in S. sonnei . This should facilitate research on this bacterium and vaccine development.

The Effect of the plant Adiantum philippense Extracts on Biofilms Formation and Adhesion to Shigella flexneri: A Predictive Modelling Approach

In the quest for novel bioactive metabolites, which can also be used as therapeutic agents, Adiantum philippense (A. philippense), an ethnomedically important fern, has become a fascinating herb. In this study, the predictive mathematical modelling of A. philippense crude extract was tested against Shigella flexneri, a common food pathogen for its phytochemical constituents, antagonistic ability, and effect on bacterial adhesion and biofilm formation was calculated. Various kinetics models such as Von Bertalanffy, Baranyi-Roberts, modified Schnute, Modified Richards, Modified Gompertz, Modified Logistics and latest Huang were used to get values for the above kinetic constants or parameters. modified Gompertz of the entire model was found to be the best model with the highest adjusted R2 value and lowest RMSE value. The accuracy and bias factors values were close to unity (1.0). The maximum specific growth rate (mmax (h-1) for S. flexneri treatment with A. philippense extract was significantly much lower (p<0.05) with a value of 0.292 (95% confidence interval of 0.254 to 0.331) compared to control with a value of 0.540 (95% confidence interval of 0.481 to 0.599) indicating potential biofilm inhibition.

Shigellosis and Development of Multiple Antimicrobial Resistance Mechanisms of Shigella spp.

Shigellosis is a serious public health issue. Millions of people suffer from this deadly food and water borne disease each year. The main manifestations of affected persons are bloody diarrhea with excessive dehydration. The causative agent of this disease is the bacteria Shigella spp. which has four serogroups. Though Shigella flexneri and Shigella dysenteriae are the dominant serogroups in developing countries, reports of other serogroups, namely Shigella boydii and Shigella sonnei, in the food contaminations are available. There are seasonal variations of Shigella infection throughout the world. In Asian subcontinent, monsoon and post monsoon times are the ideal for infection. The transmission of the bacteria in human is usually caused by feco-oral route or by contaminated food and water. There are several groups of antibiotics like foscomycin, macrolide, amiglycoside, tetracycline etc. which were used before. But they are now become useless as Shigella spp. is getting resistant against those drugs. The quinolone groups of antibiotics like ciprofloxacin, ofloxacin, norflxacin, ceftriaxone etc. are the important drugs for the cure of the disease shigellosis but prevalence of drug resistant strains of Shigella spp. against those drugs are a great concern nowadays. The occurrence of plasmid mediated quinolone resistance genes (PMQR), efflux pump proteins and effective mutations at drug binding region of gyrA etc. are the major mechanisms for the development of drug resistance.

Phenolic Compound Content and the Antioxidant and Antimicrobial Activity of Wild Blueberries (Vaccinium stenophyllum Steud.) Fruits Extracts during Ripening

The genus Vaccinium contains about 400 species distributed worldwide, but only a few species and varieties have had their phenolic composition and biological activity documented. In this study, phenols, flavonoids, anthocyanins, antioxidant and antimicrobial activity of methanolic extracts of Vaccinium stenophyllum Steud. fruits: 1-totally immature, 2-immature, 3-immature/close to maturity and 4-mature, were determined using UV-Visible spectrometry and HPTLC. The totally immature fruit extract showed the highest content of total phenols (19.153 ± 0.175 mg GAE/g DW), chlorogenic acid (20.867 ± 0.240 mg CAE/g DW), and the highest antioxidant activity by ABTS●+ (196.761 ± 0.641 µM TE/g DW) and DPPH● (146.580 ± 6.466 µM TE/g DW). Immature, immature/close to maturity and mature fruits extracts, exhibited the lowest MIC (9.37 mg/mL) and MBC (18.75 mg/mL) against Escherichia coli, Salmonella choleraesuis, and Shigella flexneri. The mature fruits extract exhibited the highest content of total anthocyanins (0.141 ± 0.004 mg CE/g DW) and cyanidin-3-glucoside (19.230 ± 0.309 mg CGE/g DW). The content of phenols, flavonoids and anthocyanins was higher than that reported for other Vaccinium species. These results showed the relevance of Vaccinium stenophyllum Steud. for breeding purposes to enhance the bioactivity of cultivars, or as a source of natural additives for the food industry, among others.

Linkage of Whole Genome Sequencing, Epidemiological, and Clinical Data to Understand the Genetic Diversity and Clinical Outcomes of Shigella flexneri among Men Who Have Sex with Men in England

Within the last 2 decades there have been an increasing number of Shigella spp. outbreaks among men who have sex with men (MSM) worldwide. In 2015, Public Health England (PHE) introduced routine whole genome sequencing (WGS) for the national surveillance of Shigella spp.

The evolutionary history of Shigella flexneri serotype 6 in Asia

Shigella flexneri serotype 6 is an understudied cause of diarrhoeal diseases in developing countries, and has been proposed as one of the major targets for vaccine development against shigellosis. Despite being named as S. flexneri , Shigella flexneri serotype 6 is phylogenetically distinct from other S. flexneri serotypes and more closely related to S. boydii . This unique phylogenetic relationship and its low sampling frequency have hampered genomic research on this pathogen. Herein, by utilizing whole genome sequencing (WGS) and analyses of Shigella flexneri serotype 6 collected from epidemiological studies (1987–2013) in four Asian countries, we revealed its population structure and evolutionary history in the region. Phylogenetic analyses supported the delineation of Asian Shigella flexneri serotype 6 into two phylogenetic groups (PG-1 and −2). Notably, temporal phylogenetic approaches showed that extant Asian S. flexneri serotype 6 could be traced back to an inferred common ancestor arising in the 18th century. The dominant lineage PG-1 likely emerged in the 1970s, which coincided with the times to most recent common ancestors (tMRCAs) inferred from other major Southeast Asian S. flexneri serotypes. Similar to other S. flexneri serotypes in the same period in Asia, genomic analyses showed that resistance to first-generation antimicrobials was widespread, while resistance to more recent first-line antimicrobials was rare. These data also showed a number of gene inactivation and gene loss events, particularly on genes related to metabolism and synthesis of cellular appendages, emphasizing the continuing role of reductive evolution in the adaptation of the pathogen to an intracellular lifestyle. Together, our findings reveal insights into the genomic evolution of the understudied Shigella flexneri serotype 6, providing a new piece in the puzzle of Shigella epidemiology and evolution.

Cholesterol-lowering Potential and Exopolysaccharide Biosynthesis of Lactobacillus spp. isolated from Human Milk

Recent research generated information that human milk is not only a valuable source of nutrition, but it also provides a complex microbial community, containing especially Lactobacillus species - the major components of a great number of commercial probiotics. New findings on potential applications of Lactobacillus species revealed that these bacteria have abilities to produce anti-microbial exopolysaccharides (EPS) and to reduce cholesterol in culture broth. In this study, we successfully isolated and screened for Lactobacillus bacteria from human milk samples, and finally obtained four strains, including L. plantarum BM7.13, L. plantarum BM29.7, L. acidophilus BM10.8 and L. rhamnosus BM30.4. Researching the probiotic activities of these strains showed that all strains were tolerant to the low pH (3.0) and 0.3% bile salts. Characterization of the probiotic properties indicated that all selected Lactobacillus isolates had ESP (125-326 mg/L) and exhibited strong antimicrobial activities against pathogenic microbes, such as Escherichia coli, Staphylococcus aureus, Shigella flexneri and Salmonella typhimurium. Our results also indicated that all strains displayed cholesterol assimilation capabilities in culture broth with the maximum figure recorded for L. plantarum BM7.13.