Vaginal Infections during Pregnancy Increase Breast Milk Microbiome Alpha Diversity and Alter Taxonomic Composition.

We previously reported that breast milk from women with (W) or without (WO) vaginal yeast infection during pregnancy differs in its immunological and antimicrobial properties, especially against pathogenic vaginal Candida sp.. Here, we investigated the differences in microbiota profiles of breast milk from these groups. Seventy-two breast milk samples were collected from lactating mothers (W, n=37; WO, n=35). The DNA of bacteria was extracted from each breast milk sample for microbiota profiling by 16S rRNA gene sequencing. Breast milk from the W-group exhibited higher alpha diversity than that from the WO-group across different taxonomic levels of class (P=0.015), order (P=0.011), family (P=0.020), and genus (P=0.030). Compositional differences between groups as determined via beta diversity showed marginal differences at taxonomic levels of phylum (P=0.087), family (P=0.064), and genus (P=0.067). The W-group showed higher abundances of families Moraxellaceae (P=0.010) and Xanthomonadaceae (P=0.008), and their genera Acinetobacter (P=0.015), Enhydrobacter (P=0.015), and Stenotrophomonas (P=0.007). Meanwhile, the WO-group showed higher abundances of genus Staphylococcus (P=0.046) and species Streptococcus infantis (P=0.025). This study shows that, although breast milk composition is affected by vaginal infection during pregnancy, this may not pose a threat to infant growth and development.

Breast milk from healthy women has higher anti-Candida properties than women with vaginal infections during pregnancy.

The aim of this study was to investigate the different immunological and antimicrobial properties of breast milk from women with (W) or without (WO) vaginal yeast infections during pregnancy in 85 lactating women (W, n = 43; WO, n = 42). Concentrations of IL-10, IgA, IgM, IgG, EGF, and TGF-α were similar in both groups. However, breast milk of women aged below 31 years old from the W-group showed higher concentration of EGF than the WO-group (p = 0.031). Breast milk from WO-group exhibited higher anti-Candida properties than W-group, both via growth inhibition and aggregation of yeast cells (p < 0.001). Correlation analysis showed that breast milk concentration of TGF-α positively correlated with concentrations of IL-10 (p = 0.001) and IgA (p = 0.021) in the W-group. Data from our present study shows that although breast milk from women with vaginal infections during pregnancy may not sufficiently hinder Candida growth, other immuno-modulatory bioactives may substitute for such a protective effect.

Sex-divergent expression of cytochrome P450 and SIRTUIN 1-7 proteins in toxicity evaluation of a benzimidazole-derived epigenetic modulator in mice.

Efforts in precision medicine to combat aberrant epigenome have led to the development of epigenetic targeting drugs. We have previously reported the capability of the BZD9L1 epigenetic modulator to impede colorectal tumour growth in vitro and in vivo through sirtuin (SIRT) inhibition. Although most benzimidazole derivatives are commonly less toxic, their effects on SIRTs and cytochrome P450 (CYP) regulations have not been explored alongside toxicity assessments. SIRTs are histone deacetylases that are crucial in maintaining metabolic homeostasis, whereas CYP is essential in drug metabolism. This study aims to determine the toxicology profile of BZD9L1 through oral acute and repeated dose toxicity evaluations, along with molecular analyses of SIRT, CYP and relevant toxicity markers through western blot and quantitative polymerase chain reaction (qPCR). BZD9L1 demonstrated no sign of acute toxicity at the limit dose (2000 mg/kg). The 28-day toxicity study highlighted the tolerability of repeated dose administration without adverse effects. BZD9L1 showed a sex-divergent regulation of hepatic SIRT1-7, CYP2A5 and CYP2D proteins. Furthermore, BZD9L1 did not induce the expression of organ injury proteins or alter the gene expression of cellular function indicators in mouse liver and kidneys, hence demonstrating, at least in part, the safety of BZD9L1 in short-term evaluations. The present study cautions for personalised strategies when employing benzimidazole-derived epigenetic therapeutics.

Probiotic Bifidobacterium lactis Probio-M8 treated and prevented acute RTI, reduced antibiotic use and hospital stay in hospitalized young children: a randomized, double-blind, placebo-controlled study.

PURPOSE: The development of probiotics has seen tremendous growth over the years, with health benefits ranging from gut health to respiratory. We thus aimed to investigate the effects of probiotic Bifidobacterium lactis Probio-M8 (2 × 1010 log CFU/day) against acute respiratory tract infections (RTI), use of antibiotics, hospitalization period and elucidate the possible mechanisms of action in hospitalized young children. METHOD: A prospective, randomized, double-blind and placebo-controlled study was performed in RTI-hospitalized children. Patients were randomized to either the probiotic (n = 60, mean age 13.81 ± 0.90 months) or placebo (n = 60, mean age 12.11 ± 0.73 months) which were administered upon admission, continued during hospitalization and 4-week post-discharged. RTI and gut health parameters were assessed at these time points using validated questionnaires while concentrations of inflammatory cytokines were assessed via oral swabs. RESULTS: Probio-M8 reduced the duration of nasal, pharyngeal and general flu-like symptoms compared to the placebo during the hospitalization period and 4-week post-discharged (P < 0.05) as compared to the placebo, with a more prevalent effect against lower respiratory tract infections (LRTI). Probio-M8 reduced prescription of antibiotic (P = 0.037), prevented new prescription of antibiotic in non-prescribed patients (P = 0.024) and reduced hospitalization period in antibiotic-prescribed patients (P = 0.004) as compared to the placebo. Oral cytokine levels of TNF-α decreased in the Probio-M8 group (P = 0.001) accompanied by increased in IL-10 (P = 0.018) over 4-week post-discharged, while the placebo group did not exhibit such an effect. Increased IL-10 in the Probio-M8 group was correlated with decreased body ache (r = - 0.296, P = 0.001), headache (r = - 0.295, P = 0.001) and pain during swallow (r = - 0.235, P = 0.010). CONCLUSION: Data from our present study show that B. lactis Probio-M8 could be a potential natural and non-drug strategy for the management of RTI in young children in a safe manner. CLINICAL TRIAL REGISTRATION: Clinical studies (Approval No. USM/JEPeM/19030177) were registered at ClinicalTrials.gov (Identifier No. NCT04122495) on September 30, 2019.

Methadone, Buprenorphine, and Clonidine Attenuate Mitragynine Withdrawal in Rats.

Background: Kratom or Mitragyna speciosa Korth has been widely used to relieve the severity of opioid withdrawal in natural settings. However, several studies have reported that kratom may by itself cause dependence following chronic consumption. Yet, there is currently no formal treatment for kratom dependence. Mitragynine, is the major psychoactive alkaloid in kratom. Chronic mitragynine treatment can cause addiction-like symptoms in rodent models including withdrawal behaviour. In this study we assessed whether the prescription drugs, methadone, buprenorphine and clonidine, could mitigate mitragynine withdrawal effects. In order to assess treatment safety, we also evaluated hematological, biochemical and histopathological treatment effects. Methods: We induced mitragynine withdrawal behaviour in a chronic treatment paradigm in rats. Methadone (1.0 mg/kg), buprenorphine (0.8 mg/kg) and clonidine (0.1 mg/kg) were i.p. administered over four days during mitragynine withdrawal. These treatments were stopped and withdrawal sign assessment continued. Thereafter, toxicological profiles of the treatments were evaluated in the blood and in organs. Results: Chronic mitragynine treatment caused significant withdrawal behaviour lasting at least 5 days. Methadone, buprenorphine, as well as clonidine treatments significantly attenuated these withdrawal signs. No major effects on blood or organ toxicity were observed. Conclusion: These data suggest that the already available prescription medications methadone, buprenorphine, and clonidine are capable to alleviate mitragynine withdrawal signs rats. This may suggest them as treatment options also for problematic mitragynine/kratom use in humans.

Mitragynine Attenuates Morphine Withdrawal Effects in Rats-A Comparison With Methadone and Buprenorphine.

BACKGROUND: Opiate addiction is a major health problem in many countries. A crucial component of the medical treatment is the management of highly aversive opiate withdrawal signs, which may otherwise lead to resumption of drug taking. In a medication-assisted treatment (MAT), methadone and buprenorphine have been implemented as substitution drugs. Despite MAT effectiveness, there are still limitations and side effects of using methadone and buprenorphine. Thus, other alternative therapies with less side effects, overdosing, and co-morbidities are desired. One of the potential pharmacotherapies may involve kratom's major indole alkaloid, mitragynine, since kratom (Mitragyna speciosa Korth.) preparations have been reported to alleviate opiate withdrawal signs in self-treatment in Malaysian opiate addicts. METHODS: Based on the morphine withdrawal model, rats were morphine treated with increasing doses from 10 to 50 mg/kg twice daily over a period of 6 days. The treatment was discontinued on day 7 in order to induce a spontaneous morphine abstinence. The withdrawal signs were measured daily after 24 h of the last morphine administration over a period of 28 abstinence days. In rats that developed withdrawal signs, a drug replacement treatment was given using mitragynine, methadone, or buprenorphine and the global withdrawal score was evaluated. RESULTS: The morphine withdrawal model induced profound withdrawal signs for 16 days. Mitragynine (5-30 mg/kg; i.p.) was able to attenuate acute withdrawal signs in morphine dependent rats. On the other hand, smaller doses of methadone (0.5-2 mg/kg; i.p.) and buprenorphine (0.4-1.6 mg/kg; i.p.) were necessary to mitigate these effects. CONCLUSIONS: These data suggest that mitragynine may be a potential drug candidate for opiate withdrawal treatment.

Methadone hydrochloride and leukemia cells: Effects on cell viability, DNA fragmentation and apoptotic proteins expression level.

Leukemia is a type of blood cancer where abnormal and immature leucocytes are produced in the bone marrow. Methadone hydrochloride is a man-made drug that is commonly used in the maintenance treatment for drug addiction. The objective of this research was to determine the cytotoxic activity and apoptotic effects of methadone hydrochloride treatment towards two leukemia cell lines which are CCRF-CEM and HL-60. CCRF-CEM and HL-60 cells were treated with methadone hydrochloride for 24 and 48 hours to determine the cytotoxic activity. IC50 at 24 hours obtained for CCRF-CEM was 121.6µmol/L while IC50 for HL-60 cells was 97.18µmol/L. Result obtained from DNA fragmentation assay showed no characteristic DNA ladder pattern in CCRF-CEM leukemia cells treated with methadone hydrochloride. Characteristics DNA ladder pattern was observed in methadone hydrochloride treated HL-60 cells. Formation of comets was seen in methadone hydrochloride treated CCRF-CEM and HL-60 cells with varying degree of DNA damage. The comets formed by methadone hydrochloride treated HL-60 cells were more prominent as compared to methadone-treated CCRF-CEM cells. The expression of apoptotic-related proteins in methadone-treated CCRF-CEM and HL-60 cells were checked by incubating the cell lysate with Raybio® Human Apoptosis Antibody Array. Significant alterations in expression level of apoptosis-related proteins in methadone hydrochloride treated CCRF-CEM cells were found involving upregulation of caspase-8 expression and downregulation of survivin expression. Methadone hydrochloride induced apoptosis in HL-60 cells involved upregulation of Bid and caspase-8 expression and downregulation of Bcl-2, p21 and survivin expression.

Lactobacillus plantarum DR7 Reduces Cholesterol via Phosphorylation of AMPK That Down-regulated the mRNA Expression of HMG-CoA Reductase.

Hypercholesterolemia is one of the primary risk factors for cardiovascular diseases. The use of lactobacilli probiotics to reduce blood cholesterol levels have been extensively reported. However, more information is needed to evaluate the possible mechanisms involved and to identify possible targets for further therapeutic development. In this study, strains of lactobacilli were screened based on the ability to assimilate cholesterol, and prevention of cholesterol accumulation in hepatic (HepG2) and intestinal (HT-29) cells. Cell free supernatant (CFS) from Lactobacillus plantarum DR7 showed a higher ability to assimilate cholesterol, reduction in cholesterol accumulation in both HepG2 and HT-29 cells, accompanied by reduced mRNA expression of HMG-CoA reductase (HMGCR) in HepG2 (p<0.05), compared to other lactobacilli. The reduction of HMGCR expression was also diminished in the presence of an AMPK inhibitor (Compound C), suggesting that L. plantarum DR7 exerted its effect via the AMPK pathway, typically via the phosphorylation of AMPK instead of the AMPK mRNA expression in HepG2 (p<0.05). Altogether, our present study illustrated that lactobacilli could exert cholesterol lowering properties along the AMPK pathway, specifically via phosphorylation of AMPK that led to reduced expression of HMGCR.

Isolation and identification of salmonella from curry samples and its sensitivity to commercial antibiotics and aqueous extracts of Camelia sinensis (L.) and Trachyspermum ammi (L.).

OBJECTIVE: To isolate Salmonella from curry samples and to evaluate the drug sensitivity of the food-borne Salmonella and its susceptibility to specific plant extracts. METHODS: Salmonella was isolated from the curry samples by standard microbiological methods and was confirmed by biochemical tests. The antibiotic susceptibility test was conducted by disc diffusion method using commercially available antibiotics such as ampicillin, tetracycline, chloramphenicol, kanamycin, and penicillin. In addition, the susceptibility of the food-borne Salmonella was also evaluated against the aqueous extracts of Camelia sinensis (L.) Theaceae (tea leaves) and the Trachyspermum ammi (L.) Apiaceae ( ajwain or omum seeds). RESULTS: Out of fifty curry samples, only seven samples were identified to have Salmonella contamination. The Salmonella isolates showed a significant drug resistance pattern except for kanamycin. The plant extracts showed a considerable antibacterial activity against the isolates, indicating the presence of antimicrobial principle which can be exploited after complete pharmacological investigations. CONCLUSIONS: The present study demonstrates the occurrence of Salmonella in the curry samples, and shows significant drug resistance against most of the commercially available antibiotics, except kanamycin. Antimicrobial effect of the plant extracts against the food-bone Salmonella suggests that dietary including medicinal herbs would be one strategy to manage food borne pathogens.

Chemical Composition and Biological Activity of the Centipeda minima (Asteraceae).

The antimicrobial activity of the Centipeda minima L. (Asteraceae) extract was evaluated against seven microorganisms using the disc diffusion method. The extract showed a broad spectrum of antimicrobial activity against all the tested bacterial strains, especially Enterobacter aerogenes, Klebsiella pneumonia, Staphylococcus aureus and Yersinia enterocolitica. The chemical composition of the extract obtained from Centipeda minima was analysed by GC-MS. Twentythree compounds, constituting about 84.75 % of the total extract, were identified. The main constituents were palmitic acid (7.47%), (Z,Z)-9-,12-octadecatrienoic acid (6.52%), (Z,Z,Z)-9-,12-octadecatrienoic acid (7.01%), phytol (7.01%), naptho[2.3-b]furan-2-(3H)-on (6.21%), 1-(1,2,3,4,7,7a-hexahydro-1,4,4,5-tetramethyl-1,3a-ethano-3aH-inden-6-yl)etanon (7.95%), 1,3,5-tri-tertbutyl- benzene (4.52%), (3Z)-2-methyl-3-octen-2-ol (5.95%) and artemisia ketone (4.98%). The extract was also tested against brine shrimp for toxicity. There was no significant toxicity as it only recorded a LC50 value of 4.92 mg/ml. The study shows that the extract is a good antimicrobial agent with potential applications in public health against diseases.