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Background: Beta-thalassaemia (β-thalassaemia) major patients are severely anaemic and require life-long blood transfusions for survival. These patients require iron-chelation therapy as a result of iron overload due to the monthly blood transfusions. The iron over load can cause oxidative damage and pro-inflammation and therefore, hasten mortality. Thus, regular monitoring of the oxidative stress and pro-inflammation status may be useful in these patients.

Aeromonads are ubiquitous in aquatic environments and have been implicated in fish and human infections. In this study, we isolated, studied antimicrobial susceptibility patterns and screened the existence of 15 virulence genes in aeromonads from two famously consumed fish species—seven marine Tiger Grouper (Epinephelus fuscoguttatus) and eight freshwater Marble Goby (Oxyeleotris marmoratus) from the aquaculture hatchery in Sabah, Malaysia. A total of 30 aeromonads (17 A. caviae, 9 A. rivuli, 4 A. dhakensis) were identified using PCR targeting GCAT gene, rpoD‐restriction fragment length polymorphism and multi‐locus phylogenetic analysis. All 30 strains were resistant to amoxicillin and cephalothin and five strains were multidrug‐resistant. Nine virulence genes (lip, ela, eno, fla, aerA, hylA, dam, alt and ser) present in A. dhakensis, suggesting the virulence potential of this species as a fish pathogen. This study offers as a baseline for future studies in monitoring and managing these two fish in aquaculture industry. [ABSTRACT FROM AUTHOR]

Abstract: Staphylococcus aureus is able to form multilayer biofilms embedded within a glycocalyx or slime layer. Biofilm formation poses food contamination risks and can subsequently increase the risk of food poisoning. Identification of food‐related S. aureus strains will provide additional data on staphylococcal food poisoning involved in biofilm formation. A total of 52 S. aureus strains isolated from sushi and sashimi was investigated to study their ability for biofilm formation using crystal violet staining. The presence of accessory gene regulator (agr) groups and 15 adhesion genes was screened and their associations in biofilm formation were studied. All 52 S. aureus strains showed biofilm production on the tested hydrophobic surface with 44% (23/52) strains classified as strong, 33% (17/52) as moderate, and 23% (12/52) as weak biofilm producers. The frequency of agr‐positive strains was 71% (agr group 1 = 21 strains; agr group 2 = 2 strains; agr group 3 = 12 strains; agr group 4 = 2 strains) whereas agr‐negative strains were 29% (15/52). Twelve adhesion genes were detected and 98% of the S. aureus strains carried at least one adhesion gene. The ebps was significantly (p < .05) associated with strong biofilm producing strains. In addition, eno, clfA, icaAD, sasG, fnbB, cna, and sasC were significantly higher in the agr‐positive group compared to the agr‐negative group. The results of this study suggest that the presence of ebps, eno, clfA, icaAD, sasG, fnbB, cna, and sasC may play an important role in enhancing the stage of biofilm‐related infections and warrants further investigation. Practical Application: This work contributes to the knowledge on the biofilm formation and the distribution of agr groups in S. aureus strains as well as microbial surface components in recognizing adherence matrix molecules of organisms isolated from ready‐to‐eat sushi and sashimi. The findings provide valuable information to further study the roles of specific genes in causing biofilm‐related infections. [ABSTRACT FROM AUTHOR]

Homozygosity for the α-thalassaemia Southeast Asian (α-SEA) and Filipino β0-thalassaemia (β-FIL) deletions can cause serious complications leading to foetal death or life-long blood transfusions. A rapid and accurate molecular detection assay is essential in populations where the deletions are common. In this study, gap-polymerase chain reaction (PCR) with high resolution melting (HRM) analysis was developed to detect both the large deletions. Melting curves at 86.9 ± 0.1 °C were generated by normal individuals without the α-SEA deletion, 84.7 ± 0.1 °C by homozygous α-SEA deletion individuals and two melting curves at 84.7 ± 0.1 °C and 86.9 ± 0.1 °C by α-SEA deletion carriers. Normal individuals without the β-FIL deletion produce amplicons with a melting temperature (Tm) at 74.6 ± 0.1 °C, homozygous β-FIL individuals produce amplicons with Tm at 73.6 ± 0.1 °C and heterozygous β-FIL individuals generate two amplicons with Tm at 73.6 ± 0.1 °C and 74.6 ± 0.1 °C. Evaluation using blinded tests on 220 DNA samples showed 100% sensitivity and specificity. The developed assays are sensitive and specific for rapid molecular and prenatal diagnosis for the α-SEA and β-FIL deletions. [ABSTRACT FROM AUTHOR]

Objective: The objective of this study was to examine the species distribution, genetic relatedness, virulence gene profiles, antimicrobial sensitivities, and resistance gene distribution of clinical Aeromonas strains from Singapore and Malaysia. Methods: A total of 210 Aeromonas clinical isolates were investigated: 116 from Singapore General Hospital and 94 archived clinical isolates from University of Malaya Medical Center, Malaysia. The isolates were genetically identified based on the gcat gene screening and the partial sequences of the rpoD housekeeping gene. Genetic relatedness, distribution of 15 virulence genes and 4 beta-lactamase resistance genes, and susceptibility patterns to 11 antimicrobial agents were compared. Results: Of the 210 Aeromonas isolates, A. dhakensis–94 (45%) was the dominant species in Singapore and Malaysia. Species composition was similar and enterobacterial repetitive intergenic consensus-PCR did not show genetic relatedness between strains from the two countries. Of the 15 virulence genes, A. dhakensis and A. hydrophila harbored the most compared with other species. Different combinations of 9 virulence genes (exu, fla, lip, eno, alt, dam, hlyA, aexU, and ascV) were present in A. dhakensis, A. hydrophila, and A. veronii from both the countries. Distribution of virulence genes was species and anatomic site related. Majority (>80%) of the strains were susceptible to all antimicrobial agents tested, except amoxicillin and cephalothin. A. dhakensis strains from Malaysia significantly harbored the cphA gene compared with A. dhakensis from Singapore. Multidrug resistance was mostly detected in strains from peritoneal fluids of dialysis patients. Conclusion: This study revealed A. dhakensis as the dominant species isolated in both geographic regions, and that it carried a high number of virulence genes. It also highlights the geographic-related differences of virulence gene distribution and antimicrobial resistance profiles of clinical Aeromonas strains from Singapore and Malaysia. [ABSTRACT FROM AUTHOR]

Staphylococcus aureus is one of the leading causes of food poisoning. Its pathogenicity results from the possession of virulence genes that produce different toxins which result in self-limiting to severe illness often requiring hospitalization. In this study of 200 sushi and sashimi samples, S. aureus contamination was confirmed in 26% of the food samples. The S. aureus isolates were further characterized for virulence genes and antibiotic susceptibility. A high incidence of virulence genes was identified in 96.2% of the isolates and 20 different virulence gene profiles were confirmed. DNA amplification showed that 30.8% (16/52) of the S. aureus carried at least one SE gene which causes staphylococcal food poisoning. The most common enterotoxin gene was seg (11.5%) and the egc cluster was detected in 5.8% of the isolates. A combination of hla and hld was the most prevalent coexistence virulence genes and accounted for 59.6% of all isolates. Antibiotic resistance studies showed tetracycline resistance to be the most common at 28.8% while multi-drug resistance was found to be low at 3.8%. In conclusion, the high rate of S. aureus in the sampled sushi and sashimi indicates the need for food safety guidelines.

Homozygosity for the α-thalassaemia Southeast Asian (α-SEA) and Filipino β°-thalassaemia (β-FIL) deletions can cause serious complications leading to foetal death or life-long blood transfusions. A rapid and accurate molecular detection assay is essential in populations where the deletions are common. In this study, gap-polymerase chain reaction (PCR) with high resolution melting (HRM) analysis was developed to detect both the large deletions. Melting curves at 86.9 ± 0.1 °C were generated by normal individuals without the α-SEA deletion, 84.7 ± 0.1 °C by homozygous α-SEA deletion individuals and two melting curves at 84.7 ± 0.1 °C and 86.9 ± 0.1 °C by α-SEA deletion carriers. Normal individuals without the β-FIL deletion produce amplicons with a melting temperature (Tm) at 74.6 ± 0.1 °C, homozygous β-FIL individuals produce amplicons with Tm at 73.6 ± 0.1 °C and heterozygous β-FIL individuals generate two amplicons with Tm at 73.6 ± 0.1 °C and 74.6 ± 0.1 °C. Evaluation using blinded tests on 220 DNA samples showed 100% sensitivity and specificity. The developed assays are sensitive and specific for rapid molecular and prenatal diagnosis for the α-SEA and β-FIL deletions.

Background: Beta-thalassemia is the most common genetic disorder in Malaysia. Confirmation of the β-globin gene mutations involved in thalassemia is usually carried out by molecular analysis of DNA extracted from leukocytes in whole blood. Molecular analysis is generally carried out when affected children are around 1 - 2 years as clinical symptoms are expressed during this period. Blood taking at this age can be distressing for the child. High yield and pure DNA extracted from non-invasive sampling methods can serve as alternative samples in molecular studies for genetic diseases especially in pediatric cases., Methods: In this study, mouthwash, saliva, and buccal cytobrush samples were collected from β-thalassemia major patients who had previously been characterized using DNA extracted from peripheral blood. DNA was extracted from mouthwash, saliva, and buccal cytobrush samples using the conventional inexpensive phenol-chloroform method and was measured by spectrophotometry for yield and purity. Molecular characterization of β-globin gene mutations was carried out using the amplification refractory mutation system (ARMS)., Results: DNA extracted from mouthwash, saliva, and buccal cytobrush samples produced high concentration and pure DNA. The purified DNA was successfully amplified using ARMS. Results of the β-globin gene mutations using DNA from the three non-invasive samples were in 100% concordance with results from DNA extracted from peripheral blood., Conclusions: The conventional in-house developed methods for non-invasive sample collection and DNA extraction from these samples are effective and negate the use of more expensive commercial kits. In conclusion, DNA extracted from mouthwash, saliva, and buccal cytobrush samples provided sufficiently high amounts of pure DNA suitable for molecular analysis of β-thalassemia.