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Molecular Profiling and Antibiotic Resistance of Salmonella Enterica Subsp. Enterica Isolated from Indigenous Ulam and Poultry Meat

Salmonella enterica subsp. enterica formed the major group that represents nearly 60% of the salmonellae. Salmonella organisms emerged as a public health problem in many countries as salmonellosis has become the most prevalent foodborne disease worldwide. It has been estimated that approximately 1.4 million cases were reported annually in the developed nations such as USA. In Malaysia, of 8,640 cases of food poisoning reported by the Ministry of Health for the year 1999, 811 (9.4%) were due to Salmonella. The purpose of this study was to characterize and study Salmonella enterica subsp. enterica (S. enterica) using multiple antimicrobial resistance and several molecular typing methods including plasmid profiling, PCR-RFLP, RAPD, ERIC-PCR and Multiplex PCR on antibiotic resistant gene. The isolate were recovered from poultry meat (55), four types of indigenous vegetables namely ‘selom’ (Oenanthe stolonifera) (59), ‘pegaga’ (Centella asiatica) (20), ‘kesum’ (Polygonum minus) (41), ‘kangkong’ (Ipomoea aquatica) (14) and processed food (11).Genomic DNA of the 200 S. enterica isolates belonging to 43 different serovars were recovered from poultry meat, various indigenous vegetables and processed food was confirmed by specific and duplex PCR targeting the iroB gene that yielded 443 bp and 606 bp amplicons. The PCR amplification of iroB gene is a rapid and reliable method for distinguishing between S. enterica and other bacterial species. Plasmids of S. enterica varied in sizes from 2 to more than 200 kb. Despite limited knowledge on their function, their presence is frequently used for strain differentiation in epidemiological studies. Plasmid profiling on the 200 S. enterica isolates demonstrated high discriminatory capability for serovars differentiation in this study that was clustered into 70 groups based on the number and pattern of the bands. One of the amplification based techniques used in this study for molecular characterization was PCR-RFLP that incorporated PCR of iroB1, iroB2 and restriction digest with BglII and AluI to determine the relatedness of bacterial strains. Results obtained showed that PCR-RFLP has excellent typeablity but low discriminatory power due to its inability to produce different banding patterns. ERIC sequences are short, highly conserved 126 bp non-coding regions found in the Enterobacteriaceae. Its location in bacterial genomes allows discrimination at the genus, species and serovars levels. RAPD is an amplification-based technique using arbitrary primers to detect changes in the DNA sequence at the sites in the genome and enable the discrimination of samples according to sources and serovars. Dendrogram of RAPD and ERIC-PCR were analyzed and comparisons made using BioNumerics gel analysis software (Applied Maths, Kortrijk, Belgium). Among the 200 isolates of S. enterica, RAPD with arbitrary primers OPAR02, OPAR17 and OPAR19 generated 47 clusters and 13 single isolates whereas ERIC-PCR with primers ERIC-1 and ERIC-2 produced 46 clusters and 12 single isolates at 60% similarity level with discriminatory index (D) of 0.9726 and 0.9606 respectively. Composite analysis of RAPD and ERIC-PCR profiling simultaneously produced 50 clusters and 18 single isolates at 60% similarity level with highest discriminatory index of 0.9824. These results demonstrated that composite analysis of RAPD (OPAR02, OPAR17 and OPAR19) together with ERIC-PCR are a better tool for differentiation and characterization of S. enterica as compared to a single method approach. The multiplex PCR targeted three different antibiotic resistance genes that was used to detect TEM, PSE-1 and cmlA/tetR genes segment encoding resistance towards ampicillin, chloramphenicol and tetracycline, respectively which could reduce labour and cost in analysis of a large number of isolates. Subsequently antimicrobial resistance was performed using disc diffusion method with a selection of 13 different antimicrobial agents. Total of 66 profiles were generated and multiple antimicrobial resistance (MAR) analysis indicated poultry meat still remains as the main reservoir for multi drug resistant Salmonella. In contrast, six isolates from the indigenous vegetables showed the highest MAR index (0.69). This might be due to animal waste fertilizer, irrigation water, contaminated container and improper handling of food by human that contributed to be the sources of Salmonella contamination of vegetables. Further investigations need to be conducted to determine if Salmonella isolates in recovered from indigenous vegetables were gaining more antimicrobial resistance. The characterization of MAR enabled the determination of antimicrobial patterns and trends in Salmonella from poultry meat and indigenous vegetables in Malaysia. As a conclusion, the results from this study could provide valuable information on the epidemiology and drug resistance trends of S. enterica, and hence contribute towards better surveillance and infection control measures as well as improved public health policy

Comparison of PCR fingerprinting techniques for the discrimination of Salmonella enterica subsp. enterica serovar Weltevreden isolated from indigenous vegetables in Malaysia.

Salmonella enterica subsp. enterica (S.) serovar Weltevreden has emerged as a public health problem in many countries. Genomic DNA of S. Weltevreden from indigenous vegetables namely 'selom' (Oenanthe stolonifera), 'pegaga' (Centella asiatica), 'kesum' (Polygonum minus) and 'kangkong' (Ipomoea aquatica) were characterized by duplex-polymerase chain reaction (duplex-PCR), multiplex-polymerase chain reaction (multiplex-PCR), random amplified polymorphic DNA (RAPD), enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The results demonstrated that a total of four clusters and three single isolates were generated from ERIC-PCR with primers ERIC-1 and ERIC-2 whereas RAPD with arbitrary primers OPAR2, OPAR17 and OPAR19 discriminated the S. Weltevreden into nine clusters and eight single isolates at a common 65% similarity level with discriminatory index (D) of 0.7443 and 0.9394 respectively. Composite analysis of banding profiles generated from RAPD-PCR and ERIC-PCR showed eight clusters and six single isolates at 65% similarity level with the highest D value that is 0.9508. On the other hand, PCR-RFLP and duplex PCR data exhibited a consistent profile for S. Weltevreden. Multiplex-PCR targeting three different antibiotic resistance genes and a common Salmonella specific gene segment produced two distinguishing profiles among the S. Weltevreden examined. These results demonstrated that the combined analysis of RAPD-PCR and ERIC-PCR is a better tool for characterizing S. Weltevreden than individual methods

Caenorhabditis elegans-based analysis of Salmonella enterica

Caenorhabditis elegans (C. elegans) have been widely used as an infection model for mammalian related pathogens with promising results. The bacterial factors required for virulence in non-mammalian host C. elegans play a role in mammalian systems. Previous reported that Salmonella found in vegetable and poultry meat could be potential health hazards to human. This study evaluated the pathogenicity of various serovars of Salmonella enterica (S. enterica) that recovered from local indigenous vegetables and poultry meat using C. elegans as a simple host model. Almost all S. enterica isolates were capable of colonizing the intestine of C. elegans, causing a significant reduction in the survival of nematodes. The colonization of Salmonella in C. elegans revealed that the ability of S. enterica in killing C. elegans correlates with its accumulation in the intestine to achieve full pathogenicity. Using this model, the virulence mechanisms of opportunistic pathogenic S. enterica were found to be not only relevant for the interactions of the bacteria with C. elegans but also with mammalian hosts including humans. Hence, C. elegans model could provide valuable insight into preliminary factors from the host that contributes to the environmental bacterial pathogenesis scenario

Gynura procumbens: An Overview of the Biological Activities

Gynura procumbens (Lour.) Merr. (Family Asteraceae) is a medicinal plant commonly found in tropical Asia countries such as China, Thailand, Indonesia, Malaysia and Vietnam. Traditionally, it is widely used in many different countries for the treatment of a wide variety of health ailments such as kidney discomfort, rheumatism, diabetes mellitus, constipation and hypertension. Based on the traditional uses of G. procumbens, it seems to possess high therapeutic potential for treatment of various diseases making it a target for pharmacological studies aiming to validate and provide scientific evidence for the traditional claims of its efficacy. Although there has been considerable progress in the research on G. procumbens, to date there is no review paper gathering the reported biological activities of G. procumbens. Hence, this review aims to provide an overview of the biological activities of G. procumbens based on reported in vitro and in vivo studies. In brief, G. procumbens has been reported to exhibit antihypertensive, cardioprotective, antihyperglycemic, fertility enhancement, anticancer, antimicrobial, antioxidant, organ protective and antiinflammatory activity. The commercial applications of G. procumbens have also been summarized in this paper based on existing patents. The data compiled illustrate that G. procumbens is a potential natural source of compounds with various pharmacological actions which can be utilised for the development of novel therapeutic agents

Virulotyping of Salmonella enterica subsp. enterica isolated from indigenous vegetables and poultry meat in Malaysia using multiplex-PCR

The increased occurrence of Salmonella occurrence in local indigenous vegetables and poultry meat can be a potential health hazards. This study is aimed to detect the prevalence of twenty different virulence factors among Salmonella enterica strains isolated from poultry and local indigenous vegetables in Malaysia via an optimized, rapid and specific multiplex PCR assay. The assay encompasses a total of 19 Salmonella pathogenicity islands genes and a quorum sensing gene (sdiA) in three multiplex reaction sets. A total of 114 Salmonella enterica isolates belonging to 38 different serovars were tested. Each isolate in under this study was found to possess up to 70% of the virulence genes tested and exhibited variable pathogenicity gene patterns. Reproducibility of the multiplex PCR assay was found to be 100% and the detection limit of the optimized multiplex PCR was tested with lowest detectable concentration of DNA 0.8 pg μl−1. This study demonstrated various Salmonella pathogenicity island virulence gene patterns even within the same serovar. This sets of multiplex PCR system provide a fast and reliable typing approach based on Salmonella pathogenicity islands, thus enabling an effective monitoring of emerging pathogenic Salmonella strains as an additional tool in Salmonella surveillance studies

Presence of antioxidative agent, Pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro- in newly isolated Streptomyces mangrovisoli sp. nov.

A novel Streptomyces, strain MUSC 149T was isolated from mangrove soil. A polyphasic approach was used to study the taxonomy of MUSC 149T, which shows a range of phylogenetic and chemotaxonomic properties consistent with those of the members of the genus Streptomyces. The diamino acid of the cell wall peptidoglycan was LL-diaminopimelic acid. The predominant menaquinones were identified as MK9(H8) and MK9(H6). Phylogenetic analysis indicated that closely related strains include Streptomyces rhizophilus NBRC 108885T (99.2 % sequence similarity), Streptomyces gramineus NBRC 107863T (98.7 %) and Streptomyces graminisoli NBRC 108883T (98.5 %). The DNA–DNA relatedness values between MUSC 149T and closely related type strains ranged from 12.4 ± 3.3 % to 27.3 ± 1.9 %. The DNA G + C content was determined to be 72.7 mol%. The extract of MUSC 149T exhibited strong antioxidant activity and chemical analysis reported identification of an antioxidant agent, Pyrrolo[1,2-a]pyrazine-1,4-dione, hexahydro-. These data showed that metabolites of MUSC 149T shall be useful as preventive agent against free-radical associated diseases. Based on the polyphasic study of MUSC 149T, the strain merits assignment to a novel species, for which the name Streptomyces mangrovisoli sp. nov. is proposed. The type strain is MUSC 149T (= MCCC 1K00699T = DSM 100438T)

Reconstructing a Reactor\u27s Multimodal Polymer Molecular Weight Distribution

Using computational fluid dynamics (CFD) and statistical methods, the molecular weight distribution (MWD) of a low-density polyethylene (LDPE) autoclave reaction will be reconstructed to predict polymer properties and optimize reaction control. LDPE reactions are highly exothermic; poor control can lead to inefficiency, hot spots, and potentially explosive decomposition. A polymer\u27s MWD significantly influences its properties, and the sensitivity of ethylene polymerization necessitates simulation techniques for safer reactor design. However, traditional polymer reactor software often assumes uniform reactant mixing, limiting its accuracy. CFD can capture spatiotemporal species gradients to create more accurate polymer reactor simulations, but it does not typically reproduce the polymer\u27s MWD explicitly. Instead, the first few moments of the distribution are tracked to determine average properties and approximate the MWD if the shape of the distribution is assumed. While convenient, this approach struggles with multimodal distributions common in industrial polymers. To address this, the polymer MWD can be divided into classes based on degree of branching, assigning each class its own MWD. Using a weighted sum on these classes\u27 MWDs will provide an overall distribution. This method will be validated in a simplified CFD reactor model and applied to a full-scale autoclave reactor, testing 5, 10, 20, and 80 polymer-class simulations. Ultimately, a multimodal MWD qualitatively consistent with plant data and literature will be achieved. This work establishes a plant-scale CFD model that incorporates detailed free-radical polymerization chemistry, offering a tool to optimize LDPE reactions efficiently and improve reactor safety

Elizabeth Warren’s DNA and What it Means for Indigenous Folx

The Texas Orato