Flexneri Strains Research Articles

Multidrug-resistant Shigella flexneri outbreak associated with a high-mortality spillover event into nonhuman primates.

Shigellosis is a gastrointestinal infection caused by species of Shigella. A large outbreak of Shigella flexneri serotype 2a occurred in Albuquerque, New Mexico (NM) between May 2021 and November 2023 that involved humans and nonhuman primates (NHP) from a local zoo. We analyzed the genomes of 202 New Mexico isolates as well as 15 closely related isolates from other states, and four from NHP. The outbreak was initially detected within men who have sex with men (MSM) but then predominantly affected people experiencing homelessness (PEH). Nearly 70% of cases were hospitalized and there was one human death. The outbreak extended into Albuquerque's BioPark Zoo, causing high morbidity and six deaths in NHPs. The NHP isolates were identical to those in the human outbreak. All isolates were multidrug-resistant, including towards fluoroquinolones, a first line treatment option which led to treatment failures in human and NHP populations. We demonstrate the transmission of this S. flexneri strain between humans and NHPs, causing fatalities in both populations. This study demonstrates the threat of antimicrobial resistant organisms to vulnerable human and primate populations and emphasizes the value of vigilant genomic surveillance within a One Health framework.

Intact and mutated Shigella diguanylate cyclases increase c-di-GMP

The intracellular human pathogen Shigella invades the colonic epithelium to cause disease. Prior to invasion, this bacterium navigates through different environments within the human body, including the stomach and the small intestine. To adapt to changing environments, Shigella uses the bacterial second messenger c-di-GMP signaling system, synthesized by diguanylate cyclases (DGCs) encoding GGDEF domains. Shigella flexneri encodes a total of 9 GGDEF or GGDEF-EAL domain enzymes in its genome, but 5 of these genes have acquired mutations that presumably inactivated the c-di-GMP synthesis activity of these enzymes. In this study, we examined individual S. flexneri DGCs for their role in c-di-GMP synthesis and pathogenesis. We individually expressed each of the 4 intact DGCs in an S. flexneri strain where these 4 DGCs had been deleted (Δ4DGC). We found that the 4 S. flexneri intact DGCs synthesize c-di-GMP at different levels in vitro and during infection of tissue-cultured cells. We also found that dgcF and dgcI expression significantly reduces invasion and plaque formation, and dgcF expression increases acid sensitivity, and that these phenotypes did not correspond with measured c-di-GMP levels. However, deletion of these 4 DGCs did not eliminate S. flexneri c-di-GMP, and we found that dgcE, dgcQ, and dgcN, which all have nonsense mutations prior to the GGDEF domain, still produce c-di-GMP. These S. flexneri degenerate DGC pseudogenes are expressed as multiple proteins, consistent with multiple start codons within the gene. We propose that both intact and degenerate DGCs contribute to S. flexneri c-di-GMP signaling.

Identification of therapeutic drug target of Shigella Flexneri serotype X through subtractive genomic approach and in-silico screening based on drug repurposing

Shigellosis, induced by Shigella flexneri, constitutes a significant health burden in developing nations, particularly impacting socioeconomically disadvantaged communities. Designated as the second most prevalent cause of diarrheal illness by the World Health Organization (WHO), it precipitates an estimated 212,000 fatalities annually. Within the spectrum of S. flexneri strains, serotype X is notably pervasive and resilient, yet its comprehensive characterization remains deficient. The present investigation endeavors to discern potential pharmacological targets and repurpose existing drug compounds against S. flexneri serotype X. Employing the framework of subtractive genomics, the study interrogates the reference genome of S. flexneri Serotype X (strain 2,002,017; UP000001884) to delineate its proteome into categories of non-homologous, non-paralogous, essential, virulent, and resistant constituents, thereby facilitating the identification of therapeutic targets. Subsequently, a screening of approximately 9000 compounds from the FDA library against the identified drug target aims to delineate efficacious agents for combating S. flexneri serotype X infections. The application of subtractive genomics methodology yields prognostic insights, unveiling non-paralogous proteins (n = 4122), non-homologues (n = 1803), essential (n = 1246), drug-like (n = 389), resistant (n = 167), alongside 42 virulent proteins within the reference proteome. This iterative process culminates in the identification of Serine O-acetyltransferase as a viable drug target. Subsequent virtual screening endeavors to unearth FDA-approved medicinal compounds capable of inhibiting Serine O-acetyltransferase. Noteworthy candidates such as DB12983, DB15085, DB16098, DB16185, and DB16262 emerge, exhibiting potential for mitigating S. flexneri Serotype X. Despite the auspicious findings, diligent scrutiny is imperative to ascertain the efficacy and safety profile of the proposed drug candidates vis-à-vis S. flexneri.

Comprehensive in vitro evaluation of Indigofera hochstetteri Baker extract: Effect of chemicals in antimicrobial, anticancer, anti-inflammatory, and anti-diabetic activities

The study aimed to analyze the pharmacological properties of medicinal plant Indigofera hochstetteri Baker extracts. Preliminary phytochemical analysis revealed a diverse range of secondary metabolites present in it. TLC analysis detected numerous phytochemicals with varying Rf values, aiding in different solvent systems. GC-MS analysis revealed the presence of 29 bioactive compounds with diverse pharmacological activities, including anti-inflammatory, antioxidant, analgesic and antimicrobial properties. Antimicrobial effect of I. hochstetteri Baker methanolic extract showed significant inhibitory effects against E. coli, E. aerogenes, S. flexneri, P. aeruginosa, S. aureus, E. faecalis, B. cereus, and fungal strain C. albicans. The methanol extract also showed significant antifungal activity by inhibiting the growth of Sclerotium rolfsii in food poisoning method. MTT assays revealed significant cytotoxic activity of methanolic extract against human leukemia HL-60 cancer cells with IC50 of 116.01 μg/mL. In apoptotic study, I. hochstetteri Baker methanolic extract showed 28.84% viable cells, 30.2% early apoptosis, 35.54% late apoptosis, and 5.86% necrosis comparatively similar with standard used. The extract showed significant anti-inflammatory effect on HRBC stabilization, and protein denaturation of BSA and egg albumin denaturation with IC50 of 193.62 μg/mL, 113.94 μg/mL respectively. In anti-diabetic assays like α-amylase, α-glucosidase, and Glucose uptake assay, I. hochstetteri extract showed good anti-diabetic effect with IC50 of 60.64 μg/mL, 169.34 μg/mL, and 205.63 μg/mL respectively. In conclusion I. hochstetteri Baker have promising bioactive metabolites with significant biological activities, it can be good substitute for the chemical drugs after successful clinical studies.

The relationship between integrons, antibiotic resistance genes and SXT resistance in Shigella flexneri strains.

To investigate the correlation between sulfamethoxazole-trimethoprim (SXT) resistance in Shigella flexneri (S.flexneri) and the presence of integrons and relevant antibiotic resistance genes. We collected 115 strains of Shigella flexneri isolated from feces of children with diarrhea in Jinan from 2012 to 2020 and determined the minimum inhibitory concentration (MIC) of SXT by Etest method. The presence of class 1, class 2, and class 3 integron genes, variable region antibiotic resistance gene cassettes, and sul1, sul2, sul3, and SXT elements were detected using polymerase chain reaction (PCR). Positive results were further analyzed by DNA sequencing and BLAST comparison. In total, the resistance rate to SXT was 60.9% among the 115 S.flexneri strains. The prevalence of class 1 and class 2 integrons were 88.7% and 87.0%, respectively, with no class 3 integrons detected. Among the strains, 13.0% carried typical class 1 integrons with variable region antibiotic resistance gene cassettes dfrA17-aadA5 and dfrV, while 85.2% carried atypical class 1 integrons with variable region antibiotic resistance gene cassette blaoxa-30-aadA1. The variable region antibiotic resistance gene cassettes of class 2 integrons were all dfrA1+sat1+aadA1. There was a statistical difference between the presence of class 1 integrons and class 2 integrons between the SXT-sensitive and resistant S.flexneri strains (χ2=22.800, χ2=16.365, P<0.01, P<0.01). Integrons carrying dfrV and dfrA1 by integrons also showed a statistical difference in SXT resistance (χ2=9.422, χ2=16.365, P<0.01, P<0.01). PCR revealed the presence of sul1 and sul2 in 13.0% and 47.0% of strains, respectively, with neither sul3 nor SXT elements detected. There was a significant difference between the presence of sul1, sul2 between the SXT-sensitive and resistant S.flexneri strains (χ2=9.588, χ2=65.445, P<0.01, P<0.01). In summary, integrons are involved in SXT resistance of S.flexneri, and dfrV, dfrA1, sul1, sul2 are closely related to SXT resistance of S.flexneri.

Genomic, transcriptomic, and phenotypic differences among archetype Shigella flexneri strains of serotypes 2a, 3a, and 6.

Given the genomic diversity between S. flexneri serotypes and the paucity of data to support serotype-specific phenotypic differences, we applied in silico and in vitro functional analyses of archetype strains of 2457T (Sf2a), J17B (Sf3a), and CH060 (Sf6). These archetype strains represent the three leading S. flexneri serotypes recommended for inclusion in multivalent vaccines. Characterizing the genomic and phenotypic variation among these clinically prevalent serotypes is an important step toward understanding serotype-specific host-pathogen interactions to optimize the efficacy of multivalent vaccines and therapeutics. This study underpins the importance for further large-scale serotype-targeted analyses.

Modeling inter- and intraspecific interaction of microorganisms in a batch culture experiment

Aim. To demonstrate the possibility of using a batch culture model to study the interaction of Salmonella Enteritidis and Salmonell Typhimurium, which are currently the leading serotypes in the etiology of salmonellosis, in an in vitro experiment both among themselves and in combination with Yersinia pseudotuberculosis and Shigella flexneri.Materials and methods. An experimental model of microorganism associations in vitro was created by co-cultivation of different microorganism strains in an LB broth. The Salmonella and Yersinia pseudotuberculosis stains stored in the museum of G.P. Somov Research Institute of Epidemiology and Microbiology at a temperature of –80 °C were used. The behavior of the associations under study was monitored for 24 h at 37 °C on a shaker. As a control, the growth curves of all the strains in monoculture were studied. The first research stage involved culturing S. Enteritidis and S. Typhimurium followed by addition of Yersinia pseudotuberculosis and Shigella flexneri strains (reference strain ATCC 12022) at the second stage.Results. In the control experiment, the growth of the tested cultures was quite similar. However, when grown in associations, S. Typhimurium outgrew S. Enteritidis. A similar study conducted with three pathogens also revealed that, when grown in monoculture, the stains exhibit no significant differences in the growth rates. However, when grown in associations, the growth of Y. pseudotuberculosis is significantly suppressed from the onset of observation during the first hour. Under these conditions, S. Enteritidis takes the dominant position.Conclusion. Our results demonstrate that batch culture can be used to study the interaction between different types of microorganisms, as well as their intraspecific relationships. We assume that the observed behavior of microorganisms can be explained, to a certain extent, by Gause’s law of competitive exclusion. This principle implies that two species competing for the same limited food source cannot coexist in one ecological niche.

Complete genome sequence of Shigella flexneri strain P099 from the gum of an adult rhesus monkey, Macaca mulatta, with gingivitis.

Shigella flexneri was associated with gingivitis, a periodontal disease, in the rhesus monkey. We report the circularized 4.8-Mbp complete genome of Shigella flexneri strain P099 isolated from the gum of an adult rhesus monkey, Macaca mulatta, with clinical symptoms of gingivitis.

Shigella Flexneri Serotypes: O-antigen Structure, Serotype Conversion, and Serotyping Methods.

Shigella flexneri is the most common cause of shigellosis in developing countries. Up to now, 23 serotypes of S. flexneri have been reported. Different serotypes result from the addition of acetyl, glucosyl, or phosphatidylethanolamine groups on the O-antigen backbone and horizontal transfer of mentioned groups can lead to serotype conversion among S. flexneri strains. Serotype conversion causes either a circulation of pre-existing serotypes or is responsible for the emergence of new serotypes. Serotype conversion plays a pivotal role in the protection and evasion of S. flexneri from the host immune response. Furthermore, spreading any new serotype can provide evolutionary advantages. Hence, information about S. flexneri O-antigen structure, serotype conversion, and serotyping methods can be helpful to understand the disease that attributes distinct serotypes in order to apply control or prevention methods in accordance with predominant serotypes over the course of time. Thus, the scope of this review is to give an overview of the serotype structures, factors involved in O-antigen modification, molecular analysis, and epidemiological evidence for the benefits of serotype conversion for S. flexneri serotypes. We are also providing a review of the typing methods.

Characterization of Shigella flexneri in northern Vietnam in 2012–2016

Introduction. Shigellosis remains a considerable public health concern in developing countries. Shigella flexneri and Shigella sonnei are prevalent worldwide and S. sonnei has been replacing S. flexneri . Gap Statement. S. flexneri still causes outbreaks of shigellosis in northern Vietnam but limited information is available on its genetic characteristics. Aim. This study aimed to characterize the genetic characteristics of S. flexneri strains from northern Vietnam. Methodology. This study used 17 isolates from eight incidents, collected in northern Vietnam between 2012 and 2016. The samples were subjected to whole genome sequencing, molecular serotyping, cluster analysis and identification of antimicrobial resistance genes. Additionally, phylogenetic analysis was performed including isolates from previous studies. Results. Clusters were identified according to spatiotemporal backgrounds. The results suggested that two incidents in Yen Bai province in 2015 and 2016 were derived from a very recent common ancestor. All isolates belonged to phylogroup (PG) 3, which was divided into two sub-lineages. Thirteen of 17 isolates, including those from the Yen Bai incidents, belonged to sub-lineage Sub-1 and were serotyped as 1a. The remaining four isolates belonged to sub-lineage Sub-2 and were the globally predominant serotype 2a. The Sub-1 S. flexneri isolates possessed the gtrI gene, which encodes the glycosyl transferase that determines serotype 1a, with bacteriophage elements in the vicinity. Conclusion. This study revealed two PG3 sub-lineages of S. flexneri in northern Vietnam, of which Sub-1 might be specific to the region.

Novel plasmids in multidrug-resistant Shigella flexneri serotypes from Pakistan

Shigellosis is the main cause of food and waterborne diarrhea and is an emerging threat to human health. The current study characterized the indigenous multidrug-resistant Shigella flexneri serotypes for their plasmid profiles and genetic diversity, to characterize the plasmid evolutionary patterns and distribution. In total, 199 identified S. flexneri isolates belonging to six different serotypes were analyzed for plasmid profiling, followed by an analysis of whole genome sequencing. All isolates of S. flexneri resistant to antibiotics harbored multiple copies of plasmids with sizes ranging from 1.25 kbp to 9.4kbp. These isolates were clustered into 22 distinct plasmid patterns, labeled as p1-p22. Among these, p1 (24%) and p10 (13%) were the predominant plasmid profiles. All S. flexneri strains were grouped into 12 clades with a 75% similarity level. Also, a significant association was observed among the plasmid patterns, p23 and p17 with the drug-resistant patterns AMC, SXT, C (19.5%) and OFX, AMC, NA, CIP (13.5%), respectively. Moreover, the most widespread plasmid patterns p4, p10, and p1 showed a significant association with the serotypes 1b (29.16%), 2b (36%), and 7a (100%), respectively. After plasmid sequence assembly and annotation analysis, a variety of small plasmids that vary in size from 973 to 6200bp were discovered. Many of these plasmids displayed high homology and coverage with plasmids from non-S. flexneri. Several novel plasmids of small size were discovered in multidrug-resistant S. flexneri. The data also showed that plasmid profile analysis is more consistent than antibiotic susceptibility pattern analysis for identifying epidemic strains of S. flexneri isolated in Pakistan.

A Pentavalent Shigella flexneri LPS-Based Vaccine Candidate Is Safe and Immunogenic in Animal Models.

A multivalent vaccine is much needed to achieve protection against predominant Shigella serotypes. Recently, we demonstrated the clinical applicability and immunogenic potential of tri-acylated S. flexneri 2a lipopolysaccharide (Ac3-S-LPS). Using a similar approach, we designed a pentavalent LPS candidate vaccine against S. flexneri 1b, 2a, 3a, 6, and Y (PLVF). In this study, we performed molecular and antigenic characterization of the vaccine candidate and its preclinical evaluation. There were no signs of acute toxicity after subcutaneous administration of PLVF in rabbits at a proposed human dose of 125 μg. No pyrogenic reactions and adverse effects associated with chronic toxicity after repeated administration of PLVF were revealed either. The immunization of mice with PLVF led to ≥16-fold increase in S. flexneri 1b-, 2a-, 3a-, 6-, and Y-specific antibodies. In a serum bactericidal antibody (SBA) assay, we registered 54%, 66%, 35%, 60%, and 60% killing of S. flexneri 1b, 2a, 3a, 6, and Y, respectively. In the guinea pig keratoconjunctivitis model, the efficacy was 50% to 75% against challenge with all five S. flexneri serotypes. These studies demonstrate that PLVF is safe, immunogenic over a wide range of doses, and provides protection against challenge with homologous S. flexneri strains, thus confirming the validity of pentavalent design of the combined vaccine.

Evaluation of the impact of Shigella virulence genes on the basis of clinical features observed in patients with shigellosis.

Shigella continues to cause significant morbidity and mortality each year, mostly in under-five children living in developing countries. We investigated the association between Shigella virulence genes and shigellosis. We randomly selected 61 S. flexneri strains isolated from patients in Bangladesh between 2009 and 2013, and evaluated the presence of 140 MDa large-virulence-plasmid (p140), and 22 virulence genes including ipaH, ial, toxin, and T3SS-related genes. We found p140 in 79% (n = 48) and ipaBCD in 90% (n = 55) strains, while seven strains were missing the p140. The prevalence of ial was 89%, ipgC and ipgE was 85%, and the prevalence for the remaining genes was < 85%. During the multivariate analysis, we found that instead of sen, the Shigella enterotoxin gene set along with several other virulence genes such as ipgA, icsB, ipgB1, spa15, and mxiC, were significantly influencing multiple clinical features relevant to shigellosis, including bloody stool, mucoid stool, and rectal straining. We believe our model will help to determine the actual disease burden by directly looking for the genetic material in clinically suggestive patients, especially when detecting the causative organisms by traditional means is difficult.

Pantoea agglomerans, Klebsiella pneumoniae, and Shigella flexneri isolated from the Cisadane River as multiresistant bacteria to copper and dyes

Copper pollution in Cisadane is a serious environmental issue that needs to be resolved immediately due to its negative impacts on river ecosystems. Bioremediation utilising indigenous bacteria offers excellent potential to restore copper‐contaminated river water. This study aimed to obtain indigenous copper‐resistant bacteria isolated from the Cisadane River as copper bioremediation agents. Bacteria from Cisadane River water samples were isolated by the spread plate method on Luria Bertani medium containing 3 mM CuSO4. Resistance was determined based on the minimum inhibitory concentration value, while copper concentration was measured using an atomic absorption spec‐ trophotometer. The results presented a total of 13 bacterial isolates with a minimum inhibitory concentration of up to 8 mM CuSO4. Sequence alignment analysis was performed on three selected copper‐resistant bacteria, i.e. isolate IrCis1, IrCis4 and IrCis13, which were identified as Pantoea agglomerans, Klebsiella pneumoniae and Shigella flexneri based on 16S rRNA, respectively. Each isolate accumulated copper at 1.19 mg, 1.34 mg and 0.92 mg/g DW of cells, with copper biosorption potentials of 73.74%, 70.17% and 67.73%, respectively. In conclusion, P. agglomerans strain IrCis1, K. pneu‐ moniae strain IrCis4 and S. flexneri strain IrCis5 isolated from the Cisadane River can be used as copper bioremediation agents.

Cluster of cases due to Shigella flexneri producing CTX-M-15 in Spain.

The aim of our study was to delineate an outbreak of gastroenteritis caused by Shigella flexneri and affecting sixteen persons between May and June 2014 in Bilbao, Spain. All patients exhibited symptoms after consuming kebab in the same kebab shop.The outbreak is described through the clinical cases, the microbiological and molecular genetic diagnosis, and the epidemiologic investigation. Minimum inhibitory concentrations for ampicillin, amoxicillin plus clavulanic acid, third and fourth generation cephalosporins, carbapenems, monobactams, aminoglycosides, fluoroquinolones, co-trimoxazole, colistin and tigecycline were measured. The S. flexneri strains were screened by PCR for TEM, SHV, CTX-M beta-lactamases and plasmidic AmpCs and aac(6')-Ib gene. Serotyping, pulsed field gel-electrophoresis, conjugation assay, plasmid sizing by S1 enzyme digestion and Southern blot hybridization were accomplished.All the S. flexneri isolates proved to be serotype 2 and produced extended-spectrum beta-lactamase (ESBL). Carbapenems, fluoroquinolones, tigecycline, colistin, and co-trimoxazole remained active antibiotics.All the strains harboured blaCTX-M-15 and blaOXA-1 genes. The strains hosted two high-molecularweight plasmids of 100 and 230 kb, respectively. According to the hybridization assay blaCTX-M-15 was located on the plasmid of 230 kb. The identical pulsotype verified the presence of outbreak.Remarkable, that one of the food handlers has travelled recently to Pakistan, where ESBL-producing Shigella strains had been reported previously. To the best of our knowledge, this is the first outbreak caused by CTX-M-15-expressing S. flexneri in Spain and as well as in Europe.

Effects of cinnamaldehyde against planktonic bacteria and biofilm formation of Shigella flexneri

Cinnamaldehyde (CA) has demonstrated anti-inflammatory, anti-tumor and anti-cancer activities; Its antimicrobial and antibiofilm actions against Shigella flexneri, on the other hand, have not been investigated. Sh. flexneri is a gram-negative foodborne pathogen that can be widely found in nature and some industrial production environments. In this current research, our aim was to examine the influences of CA on planktonic bacteria and biofilm formation. The minimum inhibitory concentration (MIC) of CA against Sh. flexneri strain was 100 μg/mL, while bacteria treated with CA showed a longer lag phase compared with the untreated control. CA effectively inactivated the Sh. flexneri in LB broth and fresh lettuce juice. CA treatment resulted in cell membrane permeability changes and dysfunction, as proven by cell membrane depolarization, decreased intracellular ATP concentration. In addition, CA was also discovered to increase the level of reactive oxygen species (ROS) in cells, and induce morphological changes in cells. Crystal violet staining showed that the biomass of biofilm was decreased significantly with CA in 24 h. Light microscopy and field emission scanning electron microscopy (FESEM) observations demonstrated decreased biofilm adhesion and destruction of biofilm architecture after treatment with CA. These findings indicated that CA acts as a natural bacteriostatic agent to control Sh. flexneri in food processing and production.

Virulence plasmid pINV as a genetic signature for Shigella flexneri phylogeny.

Shigella flexneri is a major health burden in low- and middle-income countries, where it is a leading cause of mortality associated with diarrhoea in children, and shows an increasing incidence among travellers and men having sex with men. Like all Shigella spp., S. flexneri has evolved from commensal Escherichia coli following the acquisition of a large plasmid pINV, which contains genes essential for virulence. Current sequence typing schemes of Shigella are based on combinations of chromosomal genetic loci, since pINV-encoded virulence genes are often lost during growth in the laboratory, making these elements inappropriate for sequence typing. By performing comparative analysis of pINVs from S. flexneri strains isolated from different geographical regions and belonging to different serotypes, we found that in contrast to plasmid-encoded virulence genes, plasmid maintenance genes are highly stable pINV-encoded elements. For the first time, to our knowledge, we have developed a S. flexneri plasmid multilocus sequence typing (pMLST) method based on different combinations of alleles of the vapBC and yacAB toxin–antitoxin (TA) systems, and the parAB partitioning system. This enables typing of S. flexneri pINV plasmids into distinct ‘virulence sequence types’ (vSTs). Furthermore, the phylogenies of vST alleles and bacterial host core genomes suggests an intimate co-evolution of pINV with the chromosome of its bacterial host, consistent with previous findings. This work demonstrates the potential of plasmid maintenance loci as genetic characteristics to study as well as to trace the molecular phylogenesis of S. flexneri pINV and the phylogenetic relationship of this plasmid with its bacterial host.

Development of a loop-mediated isothermal amplification assay for molecular serotyping of Shigella flexneri Serotypes 2 and Xv

This study developed and evaluated a loop-mediated isothermal amplification (LAMP) assay to simply, rapidly and accurately identify Shigella flexneri serotypes 2and Xv. The LAMP assay based on the O-antigen synthesis and modification genes of S. flexneri including gtrII, gtrX, opt and wzx was developed. Its specificity and sensitivity were evaluated with 19 serotypes of S. flexneri and 96 other Shigella species and bacterial pathogens commonly found in stool samples. This LAMP assay was completed within 20min at 61°C and could detect boiled DNA samples at concentrations as low as 1pg/μl. The S. flexneri serotype LAMP assay exhibited 100% specificity for detecting 19 S. flexneri serotypes, no 96 strains of Shigella spp. and other bacterial pathogens. This LAMP assay was used to identify S. flexneri serotypes 2 and Xv from 299 S. flexneri strains isolated in China and results were consistent with that of slide agglutination and multiplex polymerase chain reaction results for the same isolates. This LAMP assay may facilitate rapid andreliable identifying S. flexneri serotypes 2 and Xv. The present study was the first LAMP method for identifying serotypes of S. flexneri.

Evaluation of the Antibacterial Activity and Cytotoxicity of Extracts of Endophytic Bacteria Isolated from Solanum torvum Sw (Solanaceae)

The study aimed at evaluating the antibacterial and cytotoxic potential of endophytic bacteria extracts isolated from Solanum torvum Sw (Solanaceae), a plant traditionally used in the treatment of many pathologies such as diarrheal diseases. Isolation of endophytic bacteria was done on supplemented and non-supplemented nutrient media. The extracts of these endophytes were obtained after fermentation in Muller Hinton Broth (MHB). An antibacterial screening of the extracts was carried out on 04 bacterial strains (Staphylococcus aureus, Shigella flexneri, Escherichia coli and Salmonella typhimurium). The strains having demonstrated activity were retained for the determination of the minimum inhibitory concentrations (MIC). The cytotoxicity tests of the extracts were carried out on healthy macrophage cells (Raw 264.7) and healthy kidney cells (Vero). A total of 41 isolates of endophytic bacteria were isolated from S. torvum. The antibacterial screening made it possible to retain 08 extracts which demonstrated antibacterial activity on the S. aureus and S. flexneri strains with MICs ranging from 100 to 200 μg/ml. All extracts exhibited weak cytotoxic activity except for one extract. Extracts of endophytic bacteria isolated from S. torvum showed antibacterial activity and low cytotoxicity on the pathogenic strains tested. This study shows that the endophytic bacteria of S. torvum are a source of antibacterial compounds to be explored to enrich the therapeutic arsenal against bacterial infections.

Morphological and molecular identification of multi-antibiotic resistant bacteria in the wound site of diabetic ulcers

Abstract. Risqiyah W, Narulita E, Rofiqoh A, Ludfi AS, Iqbal M. 2021. Morphological and molecular identification of multi-antibiotic resistant bacteria in the wound site of diabetic ulcers. Biodiversitas 23: 663-670. The study aimed to determine the characteristics of diverse species of bacteria found in diabetic ulcers and the inhibition of antibiotics against these bacteria. The method used is Gram staining for morphological characterization, identification using 16S rRNA gene, and pour plate for antibiotic resistance test. The morphological characterization result showed that the colonies had white color, circular shape, flat elevation, entire edge, small size, and basil shape with Gram negative type of bacteria. Another result showed yellow, greenish, pellucid, and green color, irregular shape, raised elevation, undulate and lobate edge, medium size, coccus and Gram positive type of bacteria. Query cover of molecular identification showed 73-100% and 77.61-96.77% for the result of similarity identification with Alcaligenes faecalis, Shigella flexneri, Enterococcus faecalis, Proteus mirabilis, and Acinetobacter seohaensis. The identification and antibiotic resistance testing showed that the bacterial species found in diabetic ulcers was A. faecalis strain NRBC 13111, which was resistant to all tested antibiotics, except A. faecalis strain NRBC 13111 from the UB 2.3K and A. faecalis strain NRBC 13111 from the sample UB 3.4M. Enterococcus faecalis strain ATCC 19433 was resistant to ceftriaxone, ceftazidime, clindamycin and metronidazole, intermediates to cefoperazone, and sensitive to ciprofloxacin. Proteus mirabilis strain JCM 1669 was resistant to clindamycin and metronidazole, sensitive to ceftriaxone and ciprofloxacin, intermediates to ceftazidime and cefoperazone. Proteus mirabilis strain ATCC 29906, A. seohaensis strain SW-100 and S. flexneri strain ATCC 29903 were resistant to all tested antibiotics.