Therapeutic Role of Secondary Metabolites from Probiotic Strains for Ehrlich Solid Tumors in Mice.

This study aimed to screen the bioactive components in Streptococcus equinus WC1 (SE-WC1) and Limosilactobacillus reuteri GM4 (LR-GM4) and estimate the therapeutic role in Ehrlich solid tumors (EST) mice model. Forty-four male albino EST mice were assigned into 7 groups and treated daily for 2 weeks, including the EST group, the EST mice that received SE-WC1 at a low or a high dose (0.5 ml *106 or 0.5 ml *108 cfu), the EST mice that received LR-GM4 at the low or the high dose (0.5 ml *106 or 0.5 ml *108 cfu), and the EST mice that received SE-WC1 plus LR-GM4 at the low or the high dose. Tumors were harvested, weighed, examined, and used for the determination of apoptosis-related gene expression. Samples of the intestine, liver, and kidney were gathered for histological examination. The GC-MS identified 24 and 36 bioactive compounds in SE-WC1 and LR-GM4, respectively. The main compound in SE-WC1 was lupeol; however, the main compound in LR-GM4 was retinaldehyde. EST mice showed disturbances in Bcl-2, Bax, and p53 mRNA expression along with histological changes in the intestine, liver, and kidney. Administration of both bacterial strains reduced the tumor weight, alleviated the disturbances in the gene expression, and improved the histological structure of the intestine, liver, and kidney in a dose-dependent. Moreover, LR-GM4 was more effective than SE-WC1 due to its higher content of bioactive compounds. It could be concluded that these strains of probiotics are promising for the treatment of solid tumors.

Lapidilactobacillus salsurivasis sp. nov., Secundilactobacillus muriivasis sp. nov., and Streptococcus parasalivarius sp. nov., isolated from Traditional Chinese Pickle.

Four Gram-stain-positive bacterial strains (designated 475-2T, 46-6BT, 778-2T and A810-3), isolated from traditional Chinese pickle, were characterized using a polyphasic taxonomic approach. Strain 475-2T was most closely related to the type strain of Lapidilactobacillus achengensis, having 99.9% 16S rRNA gene sequence similarity, 94.1-95.1% average nucleotide identity (ANI) and 57.6% digital DNA-DNA hybridization (dDDH) values. Strain 46-6BT was most closely related to the type strain of Secundilactobacillus similis, having 99.8% 16S rRNA gene sequence similarity, 94.3-94.9% ANI and 58.9-59.2% dDDH values. Strains 778-2T and A810-3 were phylogenetically related to the type strains of Streptococcus salivarius, Streptococcus thermophilus and Streptococcus vestibularis, having 99.7-99.9% 16S rRNA gene sequence similarities, 89.1-94.4% ANI and 39.0-55.5% dDDH values. Based upon the data obtained in the present study, three novel species, Lapidilactobacillus salsurivasis sp. nov., Secundilactobacillus muriivasis sp. nov. and Streptococcus parasalivarius sp. nov., are proposed and the type strains are 475-2T (= JCM 36613T = CCTCC AB 2023258T = LMG 33412T), 46-6BT (= JCM 36612T = CCTCC AB 2023259T = LMG 33411T) and 778-2T (= JCM 36614T = CCTCC AB 2023257T = LMG 33413T), respectively.

A nisin-inducible chromosomal gene expression system based on ICE Tn5253 of Streptococcus pneumoniae, transferable among streptococci and enterococci.

The present work reports the development and validation of a chromosomal expression system in Streptococcus pneumoniae which permits gene expression under the control of Lactococcus lactis lantibiotic nisin. The system is based on the integrative and conjugative element (ICE) Tn5253 of S. pneumoniae capable of site-specific chromosomal integration and conjugal transfer to a variety of bacterial species. We constructed an insertion vector that integrates in Tn5251, an ICE contained in Tn5253, which carries the tetracycline resistance tet(M) gene. The vector contains the nisRK regulatory system operon, the L. lactis nisin inducible promoter PnisA upstream of a multiple cloning site for target DNA insertion, and is flanked by two DNA regions of Tn5251 which drive homologous recombination in ICE Tn5253. For system evaluation, the emm6.1::ha1 fusion gene was cloned and integrated into the chromosome of the Tn5253-carrying pneumococcal strain FR24 by transformation. This gene encodes a fusion protein containing the signal peptide, the 122 N-terminal and the 140 C-terminal aa of the Streptococcus pyogenes M6 surface protein joined to the HA1 subunit of the influenza virus A hemagglutinin. Quantitative RT-PCR analysis carried out on total RNA purified from nisin treated and untreated cultures showed an increase in emm6.1::ha1 transcript copy number with growing nisin concentration. The expression of M6-HA1 protein was detected by Western blot and quantified by Dot blot, while Flow cytometry analysis confirmed the presence on the pneumococcal surface. Recombinant ICE Tn5253::[nisRK]-[emm6.1::ha1] containing the nisin-inducible expression system was successfully transferred by conjugation in different streptococcal species including Streptococcus gordonii, S. pyogenes, Streptococcus agalactiae and Enterococcus faecalis. As for S. pneumoniae, the emm6.1::ha1 transcript copy number and the amount of M6-HA1 protein produced correlated with the nisin concentration used for induction in all investigated bacterial hosts. We demonstrated that this host-vector expression system is stably integrated as a single copy within the bacterial chromosome, is transferable to both transformable and non transformable bacterial species, and allows fine tuning of protein expression modulated by nisin concentration. These characteristics make our system suitable for a wide range of applications including complementation assays, physiological studies, host-pathogen interaction studies.

Isolation, identification, and antibiotic susceptibility profile of bacterial isolates from unused diapers sold in major markets in Enugu Metropolis: Clinical and policy perspectives.

Diapers, being one of the most used items for infant care, it is imperative that they are always free of contamination from microorganisms. The objective of this study is to determine the rate of bacterial contamination and the antibiotics profile of isolated bacteria from unused diapers sold in markets in Enugu Metropolis, southeast Nigeria. The study also investigated the effectiveness of the Nigerian Consumer protection laws towards maintaining the standard of care for infants and toddlers. Fifty pieces of different brands of diapers were sampled for bacterial contamination using standard bacteriological procedures. The results show that out of 50 samples analyzed, bacterial growths were identified in 9 (18%), of which five different bacterial species were isolated. The most prevalent being Escherichia coli and Staphylococcus aureus 3 (33.3%). Other bacteria isolated were Lactobacillus spp, Klebsiella spp, and Streptococcus spp 1 (11.1%). There was no statistically significant difference in the distribution of the bacterial contamination of diapers across the different brands(p>0.05). We conclude that baby diapers sold in markets in Enugu metropolis are prone to contamination with bacteria.We recommend that appropriate measures should be taken during the manufacturing process to reduce or prevent the incidence of bacterial contamination of diapers.

Les couches étant l'un des articles les plus utilisés pour les soins des nourrissons, il est impératif qu'elles soient toujours exemptes de contamination par des micro-organismes. L'objectif de cette étude est de déterminer le taux de contamination bactérienne et le profil antibiotique des bactéries isolées provenant de couches inutilisées vendues sur les marchés de la métropole d'Enugu, dans le sud-est du Nigeria. L'étude a également examiné l'efficacité des lois nigérianes sur la protection des consommateurs pour maintenir le niveau de soins pour les nourrissons et les jeunes enfants. Cinquante morceaux de couches de différentes marques ont été échantillonnés pour détecter toute contamination bactérienne en utilisant des procédures bactériologiques standard. Les résultats montrent que sur 50 échantillons analysés, des croissances bactériennes ont été identifiées dans 9 (18 %), parmi lesquels cinq espèces bactériennes différentes ont été isolées. Les plus répandus sont Escherichia coli et Staphylococcus aureus 3 (33,3 %). Les autres bactéries isolées étaient Lactobacillus spp, Klebsiella spp et Streptococcus spp 1 (11,1 %). Il n'y avait pas de différence statistiquement significative dans la répartition de la contamination bactérienne des couches entre les différentes marques (p>0,05). Nous concluons que les couches pour bébés vendues sur les marchés de la métropole d'Enugu sont sujettes à la contamination bactérienne. Nous recommandons que des mesures appropriées soient prises pendant le processus de fabrication pour réduire ou prévenir l'incidence de la contamination bactérienne des couches.

Extraction of high-molecular-weight DNA from Streptococcus spp. for nanopore sequencing in resource-limited settings.

The long-read sequencing platform MinION, developed by Oxford Nanopore Technologies, enables the sequencing of bacterial genomes in resource-limited settings, such as field conditions or low- and middle-income countries. For this purpose, protocols for extracting high-molecular-weight DNA using nonhazardous, inexpensive reagents and equipment are needed, and some methods have been developed for gram-negative bacteria. However, we found that without modification, these protocols are unsuitable for gram-positive Streptococcus spp., a major threat to fish farming and food security in low- and middle-income countries. Multiple approaches were evaluated, and the most effective was an extraction method using lysozyme, sodium dodecyl sulfate, and proteinase K for lysis of bacterial cells and magnetic beads for DNA recovery. We optimized the method to consistently achieve sufficient yields of pure high-molecular-weight DNA with minimal reagents and time and developed a version of the protocol which can be performed without a centrifuge or electrical power. The suitability of the method was verified by MinION sequencing and assembly of 12 genomes of epidemiologically diverse fish-pathogenic Streptococcus iniae and Streptococcus agalactiae isolates. The combination of effective high-molecular-weight DNA extraction and MinION sequencing enabled the discovery of a naturally occurring 15 kb low-copy number mobilizable plasmid in S. iniae, which we name pSI1. We expect that our resource-limited settings-adapted protocol for high-molecular-weight DNA extraction could be implemented successfully for similarly recalcitrant-to-lysis gram-positive bacteria, and it represents a method of choice for MinION-based disease diagnostics in low- and middle-income countries.

Prevalence and antimicrobial resistance trends among vaginal bacteria isolates from pregnant bitches.

Many of the reproductive tract infections in the bitches are caused by bacteria that can normally be present on the vaginal mucosa. These bacteria also might have an important role as the cause responsible for pregnancy loss and fetal deaths. The choice of antibiotic therapy for the pregnant animal is narrow and represents a severe problem in veterinary practice, especially due to increased antimicrobial resistance. Due to incorrect antimicrobials use in breeding kennels, the aim of the present study was to assess the occurrence of the bacterial flora isolated from the pregnant bitches and their antibiotic sensitivity. The study was carried out at the private Veterinary clinic in Novi Sad, Serbia. The vaginal swabs were taken from 60 bitches diagnosed with pregnancy and were sent to be laboratory tested. Based on the results, the most common isolated pathogens were Staphylococcus pseudintermedius (20%) and beta-hemolytic streptococci (18.33%). Furthermore, significant resistance to antibiotics from beta-lactams group was detected. It is of particular importance that antimicrobial treatment be evidence based in order to reduce the overuse of antimicrobials due to increased concern regarding antimicrobial resistance.

Recurrent streptococcal toxic shock syndrome due to distinct episodes of Streptococcus dysgalactiae and Streptococcus agalactiae septic arthritis.

Streptococcal toxic shock syndrome (STSS) is an uncommon disorder characterised by hypotension and multiorgan failure in the setting of streptococcal infection. Recurrent STSS is rare and has been due to recurrence of the same streptococcal species. Here, we present a case of a patient who developed recurrent STSS from a Streptococcus dysgalactiae right native joint septic arthritis and subsequently from a Streptococcus agalactiae left native joint septic arthritis.

Pseudocin 196, a novel lantibiotic produced by Bifidobacterium pseudocatenulatum elicits antimicrobial activity against clinically relevant pathogens.

Bacteriocins are broad or narrow-spectrum antimicrobial compounds that have received significant scientific attention due to their potential to treat infections caused by antibiotic-resistant pathogenic bacteria. The genome of Bifidobacterium pseudocatenulatum MM0196, an antimicrobial-producing, fecal isolate from a healthy pregnant woman, was shown to contain a gene cluster predicted to encode Pseudocin 196, a novel lantibiotic, in addition to proteins involved in its processing, transport and immunity. Following antimicrobial assessment against various indicator strains, protease-sensitive Pseudocin 196 was purified to homogeneity from cell-free supernatant. MALDI TOF mass spectrometry confirmed that the purified antimicrobial compound corresponds to a molecular mass of 2679 Da, which is consistent with that deduced from its genetic origin. Pseudocin 196 is classified as a lantibiotic based on its similarity to lacticin 481, a lanthionine ring-containing lantibiotic produced by Lactococcus lactis. Pseudocin 196, the first reported bacteriocin produced by a B. pseudocatenulatum species of human origin, was shown to inhibit clinically relevant pathogens, such as Clostridium spp. and Streptococcus spp. thereby highlighting the potential application of this strain as a probiotic to treat and prevent bacterial infections.

Bactofencin YH, a novel bacteriocin with high inhibitory activity against clinical Streptococcus species.

Strain Lactiplantibacillus plantarum D1 with bacteriocin producing ability was found in the intestine of Gambusia affinis. The bacteriocin was found to have high inhibitory activity against multiple Streptococcus species and several other Gram-positive and Gram-negative bacteria. Bacteriocin was purified from culture supernatant by ion-exchange chromatography, Sep-Pak C18 cartridge, and reverse-phase high-performance liquid chromatography (RP-HPLC). Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectral analysis determined that purified bacteriocin has a molecular mass of 2,731 Da. A partial N-terminal sequence KRKKHKXQIYNNGM was obtained from the Edman analysis. The N-terminal sequence was employed to search against a translation of the draft genome of strain D1. The translated full amino acid sequence of the mature peptide is as follows: NH2- KRKKHKCQIYNNGMPTGQYRWC, which has a molecular weight of 2738 Da. A BLAST search revealed that this bacteriocin was most similar to bactofencin A but differed from it with three amino acid residues. No identical peptide or protein has been previously reported, and this peptide, termed bactofencin YH, was therefore considered to be a new bacteriocin produced by Lactiplantibacillus plantarum D1.

Activity of N-Chlorotaurine against Periodontal Pathogens.

Dental plaque bacteria play an important role in the pathogenicity of periodontitis and peri-implantitis. Therefore, antimicrobial agents are one means of treatment. N-chlorotaurine (NCT) as an endogenous well-tolerated topical antiseptic could be of advantage for this purpose. Accordingly, its microbicidal activity against some dental plaque bacteria was investigated at therapeutic concentrations in vitro. In quantitative killing assays, the activity of NCT against planktonic bacteria and against biofilms grown for 48 h on implantation screws was tested. Electron microscopy was used to demonstrate the formation of biofilm and its morphological changes. The killing of planktonic bacteria of all tested species, namely Streptococcus sanguinis, Streptococcus salivarius, Streptococcus oralis, Streptococcus cristatus, Rothia aeria, and Capnocytophaga ochracea, was shown within 10-20 min by 1% NCT in 0.01 M phosphate-buffered saline at 37 °C. Bacteria grown on screws for 24 h were inactivated by 1% NCT after 15-20 min as well, but the formation of biofilm on the screws was visible in electron microscopy not before 48 h. The killing of biofilms by 1% NCT was demonstrated after 30 min (streptococci) and 40 min (R. aeria). As expected, NCT has broad activity against dental plaque bacteria as well and should be further investigated on its clinical efficacy in periodontitis and peri-implantitis.

In vitro characterization of probiotic potential of Lactobacillus plantarum CM49 against selected cattle mastitogens.

BACKGROUND: Bovine mastitis results in significant economic losses for the dairy industry globally due to milk production losses and decreased herd efficiency. This research aimed to isolate, select, and characterize indigenous lactobacilli with probiotic properties. A total of 40 lactobacilli were isolated from healthy milk samples of cattle and identified at the species level through 16S rDNA sequencing. All isolates were initially screened for antimicrobial activity, and selected isolates underwent in vitro assessment of probiotic properties. RESULTS: Among the lactobacilli isolates, varying levels of activity (9 to 19 mm) against cattle mastitogens; Stapylococcus aureus (Staph. aureus), Escherichia coli (E. coli) and Streptococcus dysgalactiae (Strep. dysgalactiae) were observed in the well diffusion assay. These isolates demonstrated auto-aggregation (ranging from 14.29 ± 0.96% to 62.11 ± 1.09%) and co-aggregate (ranging from 9.21 ± 0.14% to 55.74 ± 0.74%) with mastitogens after 2 h. Lactobacillus (Lb.) plantarum CM49 showed sensitivity to most antibiotics tested and exhibited strong inhibitory effects, with mean log10 reductions of 3.46 for Staph. aureus, 2.82 for E. coli, and 1.45 for Strep. dysgalactiae in co-culture experiments. Furthermore, Lb. plantarum CM49 significantly decreased the adhesion rate of mastitogens on the bovine mammary cell line and mouse model, demonstrating its potential effectiveness in preventing mastitis. CONCLUSION: It is concluded that Lb. plantarum CM49 has remarkable probiotic potential with activity against cattle mastitogens in the laboratory and cell culture and competitively excludes mastitogens from bovine mammary cells and ameliorates Staph. aureus-induced mastitis in mice.

Streptococcus equi subspecies equi from strangles suspected equines: molecular detection, antibiogram profiles and risk factors.

Strangles, caused by Streptococcus equi subspecies equi, is a highly infectious disease of equines causing major health issues and financial losses. The aim of the study was to detect the presence of the SeM gene in Streptococcus equi isolated from equine suspected of having strangles. A cross-sectional study design was conducted from July to December 2022 in five districts of the central Gondar zone, Ethiopia. One-hundred sixty swab samples were taken from animals that had been clinically suspected. The SeM gene was detected using polymerase chain reaction, and the antimicrobial susceptibility test was performed using the Kirby-Bauer disc diffusion method. The binary logistic regression model was employed to test for statistical significance. In 31.87% (51/160) of the samples, Streptococcus equi species were isolated, and 31.37% (16/51) of these species carried the SeM gene. There was a significant amount of tetracycline (81.5%), erythromycin (81.5%), and vancomycin (75.5%) resistance among the 16 isolates. Strangles were more likely to be present in animals who shared feed containers (AOR = 7.59; 95% CI = 1.44-39.93), drank from the same water troughs (AOR = 7.74; 95% CI = 1.44-41.01), and spent the night together (AOR = 5.97; 95% CI 1.41-25.37). The findings of this study showed that the research areas harboured Streptococcus equi subspecies equi. Sharing feed containers and water troughs were potential sources of strangles infection; thus, these containers need to be cleaned regularly.

Streptococcus equi subspecies zooepidemicus: an emergent cause of meningoencephalitis in Northeastern Brazil.

This study describes an outbreak of Streptococcus equi subspecies zooepidemicus infections that caused meningoencephalitis and bacteremia related to unpasteurized milk consumption in northeastern Brazil. Epidemiological investigations and a brief literature review were conducted. Strains with possible neurotropism had not been identified in Brazil before these cases; however, in 2023, another case of meningoencephalitis caused by Streptococcus equi sp. zooepidemicus was described, revealing the need to maintain surveillance and highlighting that these neurotropic strains continue to circulate in the environment.

Streptococcus taoyuanensis sp. nov., a Novel Species Isolated from a Patient with Bacteremia.

Streptococcus spp. are important opportunistic pathogen of bacteremia in both immunocompetent and immunosuppressed patients. A streptococcal strain, designated ST2T, was isolated from the blood specimen of a bacteremic patient. Comparative analyses of 16S rRNA, rpoB and groEL gene sequences demonstrated that the novel strain ST2T is a member of the genus Streptococcus. Based on of 16S rRNA gene sequence similarities, the type strains of Streptococcus (S.) parasanguinis (99.2%), S. ilei (98.8%), S. oralis subsp. oralis (97.6%), S. australis (97.5%) and S. sanguinis (97.5%) were the closest neighbours to strain ST2T. The housekeeping gene sequences (rpoB and groEL) similarities of strain ST2T to these closely related type strains were 80.4-97.4%, respectively. The complete draft genome of strain ST2T consisted of 2,155,906 bp with a G + C content of 42.0%. Strain ST2T has an average nucleotide identity (ANI) value of 94.1 and 81.3% with S. parasanguinis ATCC 15912T and S. ilei I-G2T, respectively. The highest in silico DNA-DNA hybridization value with respect to the closest species S. parasanguinis was 55.6%, below the species cut-off of 70% hybridization. The primary cellular fatty acids of strain ST2T were C16:0, C18:1 ω9c, C18:0 and C14:0. Based on biochemical criteria and molecular genetic evidence, it is proposed that strain ST2T be assigned to a new species of the genus Streptococcus as Streptococcus taoyuanensis sp. nov. The type strain of Streptococcus taoyuanensis is ST2T (=NBRC 115928T = BCRC 81374T) as the type strain.

Identification and Characterization of a Novel Prophage Lysin against Streptococcus dysgalactiae.

Streptococcus dysgalactiae infection can cause bovine mastitis and lead to huge economic losses for the dairy industry. The abuse of antibiotics has resulted in growing drug resistance of S. dysgalactiae, which causes hard-to-treat infections. Bacteriophage lysin, as a novel antibacterial agent, has great potential for application against drug-resistant gram-positive bacteria. However, few studies have been conducted on the prophage lysin of S. dysgalactiae. In this study, we mined a novel prophage lysin, named Lys1644, from a clinical S. dysgalactiae isolate by genome sequencing and bioinformatic analysis. Lys1644 was expressed and purified, and the lytic activity, antibacterial spectrum, optimal pH and temperature, lytic activity in milk in vitro, and synergistic bacteriostasis with antibiotics were assessed. The Lys1644 prophage lysin showed high bacteriolysis activity specifically on S. dysgalactiae, which resulted in CFU 100-fold reduction in milk. Moreover, Lys1644 maintained high activity over a wide pH range (pH 5-10) and a wide temperature range (4-42 °C). Synergistic bacteriostatic experiments showed that the combination of low-dose Lys1644 (50 µg/mL) with a subinhibitory concentration of aminoglycoside antibiotics (kanamycin or spectinomycin) can completely inhibit bacterial growth, suggesting that the combination of Lys1644 and antibiotics could be an effective therapeutic strategy against S. dysgalactiae infection.

An Autopsy Case of Streptococcus dysgalactiae subsp. equisimilis Infectious Endocarditis Accompanied by Streptococcal Toxic Shock Syndrome.

A 66-year-old woman with liver cirrhosis and hemodialysis was referred with a 1-week history of pain and rash on the left lower leg. On an examination, the patient was in shock. She was administered catecholamine support for septic shock and ampicillin/sulbactam for severe cellulitis. Streptococcus dysgalactiae subsp. equisimilis (SDSE) was isolated from the blood culture, and she was diagnosed with streptococcal toxic shock syndrome. Despite therapy, the patient died on day 7 of admission. Infective endocarditis (IE) was diagnosed during an autopsy. Clinicians should be aware that overwhelming SDSE-IE can occur even in the absence of necrotizing fasciitis, especially in immunocompromised patients.

Molecular microbiological profiling of bottled unsweetened tea beverages: A screening experiment.

To explore the potential storage and safety of drinking leftover bottled tea beverages from various manufacturers after direct drinking from bottles, we conducted a screening experiment on the growth of salivary bacteria in plastic bottles of tea. The diluted saliva samples from 10 participants were inoculated into the test bottled beverages, which resulted in bacteria, particularly former members of the genus Lactobacillus, growing in some green tea beverages with a neutral pH. In contrast, tea beverages with less bacterial growth contained Streptococcus spp., and the leftovers may be safe to store and drink again.