Multicentre case-control study of pneumococcal infections among children with pneumonia in Peninsular Malaysia (MY-Pneumo): a study protocol.

BACKGROUND: S. pneumoniae (SPN) is the most common cause of pneumonia. The disease can be effectively prevented through immunisation. Since December 2020, the Malaysian Government has included the 10-valent pneumococcal conjugate vaccine (PCV10) for all infants born on or after 1 January 2020 as part of the National Immunisation Programme (NIP). However, the epidemiology of pneumonia remains poorly understood. To fill the knowledge gap, we established a multicentre surveillance study to understand the burden of pneumococcal pneumonia among young children in Peninsular Malaysia. METHODS: MY-Pneumo is a multicentre prospective case-control study conducted in three sentinel sites located in three different states of Peninsular Malaysia - Kuala Lumpur, Pahang, and Kelantan. A cohort of at least 500 incident cases and 500 controls is enrolled beginning in October 2021 and matched for age. Cases are hospitalised children < 5 years with radiologically confirmed pneumonia, and the controls are children without any features suggestive of pneumonia. Clinical samples, including nasopharyngeal swabs (NPS) and urine, are collected according to the study protocol. Biological fluids such as blood, cerebrospinal fluid (CSF) and pleural fluid are obtained from invasive pneumonia disease (IPD) patients, if available. All children are tested for SPN using polymerase chain reaction (PCR) and pneumococcal urine antigen test (PUAT) using BinaxNow. DISCUSSION: Surveillance data, including carriage rate, serotype variations and the phylogeny data structure of SPN among young children in Malaysia during PCV implementation, will be generated from this study. Trends and patterns of pneumococcal serotypes by different regions are important for targeted public health strategies. Our data will provide baseline information for estimating the impact of PCV10 implementation and will influence policymakers' decisions regarding the upgrade from PCV10 to a higher-valency conjugate vaccine in Malaysia. TRIAL REGISTRATION: This project was registered at ClinicalTrials.gov (NCT04923035) on 2021, June 11. The study protocol was approved by the International Medical University Joint-Committee on Research & Ethics (4.15/JCM-216/2021) and the Institutional Review Board at sentinel sites (USM/JEPeM/21020190, IREC 2021-114, MREC ID No: 2021128-9769) and University of Southampton's Ethics and Research Governance (ERGo II 64844).

Potential Usefulness of Bacteriophages for the Treatment of Multidrug-Resistant Acinetobacter Infection.

Bacteriophages were discovered in early 20th century. However, the interest in bacteriophage research was reduced with the discovery of antibiotics. With the increasing number of infections due to multidrug-resistant (MDR) organisms, the potential usefulness of bacteriophages as therapeutic agents has been re-evaluated. In this review, we found that more than 30 lytic bacteriophages that infect Acinetobacter species have been characterised. These are mainly members of Caudovirales, with genome sizes ranging from 31 kb to 234 kb and G+C contents ranging from 33.5% to 45.5%. The host range can be as low as < 10% of all tested Acinetobacter strains. Fourteen published murine trials indicated positive outcomes in bacteriophage-treated groups. Only two case reports were pertaining to the use of bacteriophages in the treatment of Acinetobacter infections in humans; in both cases, the infections were resolved with bacteriophage therapy. The use of bacteriophages has been associated with reduced Acinetobacter burden in the environment, as shown in two studies. The major limitation of bacteriophage therapy is its highly selective host strain. In conclusion, the potential usefulness of bacteriophage therapy for the treatment of MDR Acinetobacter species has been documented only in limited studies and more research is needed prior to its extensive use in clinical practice.

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.

Impact of a Health Education Intervention on the Incidence of Influenza-Like Illnesses (ILI) During Hajj via Smartphone Application.

This study was aimed at demonstrating the impact of a health education intervention on reducing the incidence of influenza-like illnesses symptoms among Malaysian Hajj pilgrims. This study utilizes a quasi-experimental study in the evaluation of the impact of the intervention. Participants were recruited during Hajj orientation courses organized by private Hajj companies. Participants from two companies were assigned to an intervention group and control group, respectively. The intervention group received a smartphone-based health education intervention guided by the Health Belief Model (HBM), while the control group received a regular Hajj guide intervention smartphone application before departure to Hajj. Data on the incidence of influenza-like illnesses (ILI) were collected from participants from two Hajj companies before and after returning from Hajj. Data analysis was conducted using SPSS with descriptive analysis, and analytical tests were conducted at 5% significance level. A total of 102 pilgrims completed the study in both intervention and control groups. The incidence of ILI and Non-ILI symptoms were statistically significant when the intervention and control groups (p = 0.049) were compared. In conclusion, health education has an impact on reducing the incidence of ILI and non-ILI among Hajj pilgrims.

Gaps in antimicrobial stewardship programmes in Asia: a survey of 10 countries.

Objectives: To determine antimicrobial stewardship (AMS) programme practices in Asian secondary- and tertiary-care hospitals. Methods: AMS programme team members within 349 hospitals from 10 countries (Cambodia, India, Indonesia, Japan, Malaysia, Pakistan, the Philippines, Taiwan, Thailand and Vietnam) completed a questionnaire via a web-based survey link. The survey contained questions as to whether 12 core components deemed essential for AMS programmes were implemented. Results: Overall, 47 (13.5%) hospitals fulfilled all core AMS programme components. There was a mean positive response rate (PRR) of 85.6% for the responding countries in relation to a formal hospital leadership statement of support for AMS activities, but this was not matched by budgeted financial support for AMS activities (mean PRR 57.1%). Mean PRRs were ≥80.0% for the core AMS team comprising a physician or other leader responsible for AMS activities, a pharmacist and infection control and microbiology personnel. Most hospitals had access to a timely and reliable microbiology service (mean PRR 90.4%). Facility-specific antibiotic treatment guidelines for common infections (mean PRR 78.7%) were in place more often than pre-authorization and/or prospective audit and feedback systems (mean PRR 66.5%). In terms of AMS monitoring and reporting, PRRs of monitoring specific antibiotic use, regularly publishing AMS outcome measures, and the existence of a hospital antibiogram were 75.1%, 64.4% and 77.9%, respectively. Conclusions: Most hospitals participating in this survey did not have AMS programmes fulfilling the requirements for gold standard AMS programmes in hospital settings. Urgent action is required to address AMS funding and resourcing deficits.

Prevalence of Multidrug-Resistant and Extended-Spectrum Beta-Lactamase-Producing Shigella Species in Asia: A Systematic Review and Meta-Analysis.

Shigellosis remains one of the leading causes of morbidity and mortality worldwide and is the second leading cause of diarrheal mortality among all age groups. However, the global emergence of antimicrobial-resistant Shigella strains, limiting the choice of effective drugs for shigellosis, has become the major challenge in the treatment of Shigella infections. The aim of this systematic review and meta-analysis was to provide an updated picture of the prevalence of antimicrobial-resistant Shigella species in Asia. A comprehensive and systematic search was performed on three electronic databases (PubMed, ScienceDirect and Scopus), in which 63 eligible studies published between 2010 and 2022 were identified. From our meta-analysis of proportions using a random-effects model, the overall prevalence of Shigella spp. in Asian patients was estimated to be 8.0% (95% CI: 5.5-10.5). The pooled prevalence rates of multidrug-resistant (MDR) and extended-spectrum beta-lactamase (ESBL)-producing Shigella strains were 68.7% (95% CI: 59.9-77.5) and 23.9% (95% CI: 12.9-34.8), respectively. Concerning recommended antimicrobial drugs for Shigella, the prevalence of resistance was highest for ciprofloxacin (29.8%) and azithromycin (29.2%), followed by ceftriaxone (23.8%), in spite of their importance as first- and second-line treatments for shigellosis. In contrast, resistance to carbapenems, such as ertapenem (0.0%), imipenem (0.1%) and meropenem (0.0%), was almost non-existent among the 49 tested antibiotics. The significantly high prevalence estimation suggests that the multidrug-resistant Shigella is a pressing threat to public health worthy of careful and justified interventions. Effective antibiotic treatment strategies, which may lead to better outcomes for the control and treatment of shigellosis in Asia, are essential.

Prevalence of Multidrug-Resistant Diarrheagenic Escherichia coli in Asia: A Systematic Review and Meta-Analysis.

Diarrhea is one of the leading causes of morbidity and mortality in developing countries. Diarrheagenic Escherichia coli (DEC) is an important bacterial agent for diarrhea in infants, children, and international travelers, and accounts for more than 30% of diarrheal cases in children less than 5 years old. However, the choices of antimicrobial agents are now being limited by the ineffectiveness of many first-line drugs, in relation to the emergence of antimicrobial-resistant E. coli strains. The aim of this systematic review and meta-analysis was to provide an updated prevalence of antimicrobial-resistant DEC in Asia. A comprehensive systematic search was conducted on three electronic databases (PubMed, ScienceDirect, and Scopus), where 40 eligible studies published between 2010 and 2022 were identified. Using meta-analysis of proportions and a random-effects model, the pooled prevalence of DEC in Asian diarrheal patients was 22.8% (95% CI: 16.5-29.2). The overall prevalence of multidrug-resistant (MDR) and extended-spectrum beta-lactamase (ESBL)-producing DEC strains was estimated to be 66.3% (95% CI: 58.9-73.7) and 48.6% (95% CI: 35.1-62.1), respectively. Considering antimicrobial drugs for DEC, the resistance prevalence was highest for the penicillin class of antibiotics, where 80.9% of the DEC isolates were resistant to amoxicillin and 73.5% were resistant to ampicillin. In contrast, resistance to carbapenems such as imipenem (0.1%), ertapenem (2.6%), and meropenem (7.9%) was the lowest. The relatively high prevalence estimation signifies that the multidrug-resistant DEC is a public health threat. Effective antibiotic treatment strategies, which may lead to better outcomes for the control of E. coli infections in Asia, are necessary.

Low Seroprevalence of SARS-CoV-2 among Healthcare Workers in Malaysia during the Third COVID-19 Wave: Prospective Study with Literature Survey on Infection Prevention and Control Measures.

Healthcare workers (HCWs) are at greater risk for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. This serology surveillance study aimed to investigate the prevalence of SARS-CoV-2 antibodies among the HCWs who were asymptomatic during the third wave of COVID-19 in Malaysia. HCWs from the Universiti Sains Malaysia (USM) Health Campus were prospectively recruited between August 2020 and March 2021 on a voluntary basis. Data on socio-demographics, possible risk factors and travel history were recorded. Serological diagnoses from serum samples were examined for total antibodies against SARS-CoV-2 using an immunoassay kit. A literature survey was performed on the compliance with infection and prevention control (IPC) practices for COVID-19 among HCWs. The majority of the total 617 HCWs participating in this study were nurses (64.3%, n = 397), followed by health attendants (20.9%, n = 129), medical doctors (9.6%, n = 59) and others (6.3%, n = 39). Of those, 28.2% (n = 174) claimed to have exposure to COVID-19 cases, including history of close contact and casual contact with infected patients. Most importantly, all serum samples were found to be non-reactive to SARS-CoV-2, although nearly half (40.0%, n = 246) of the HCWs had been involved directly in the management of acute respiratory illness cases. A proportion of 12.7% (n = 78) of the HCWs reported having underlying health problems, such as diabetes mellitus, hypertension and hyperlipidemia. Despite the presence of medical and sociological risks associated with SARS-CoV-2 infections, the current study found zero prevalence of antibodies against SARS-CoV-2 among the HCWs of USM. Based on the literature survey, the vast majority of Malaysian HCWs demonstrated good IPC practices during the pandemic (average percentage ranged between 92.2% and 99.8%). High compliance with IPC measures may have led to the low seroprevalence of SARS-CoV-2 among the HCWs.

Structural Insights into Substrate Binding and Antibiotic Inhibition of Enterobacterial Penicillin-Binding Protein 6.

Shigella sonnei remains the second most common cause of shigellosis in young children and is now increasingly dominant across developing countries. The global emergence of drug resistance has become a main burden in the treatment of S. sonnei infections and ß-lactam antibiotics, such as pivmecillinam and ceftriaxone, are recommended to be used as second-line treatment. They work by inhibiting the biosynthesis of the peptidoglycan layer of bacterial cell walls, in which the final transpeptidation step is facilitated by penicillin-binding proteins (PBPs). In this study, using protein homology modelling, we modelled the structure of PBP6 from S. sonnei and comprehensively examined the molecular interactions between PBP6 and its pentapeptide substrate and two antibiotic inhibitors. The docked complex of S. sonnei PBP6 with pentapeptides showed that the substrate bound to the active site groove of the DD-carboxypeptidase domain, via hydrogen bonding interactions with the residues S79, V80, Q101, G144, D146 and R240, in close proximity to the catalytic nucleophile S36 for the nucleophilic attack. Two residues, R240 and T208, were found to be important in ligand recognition and binding, where they formed strong hydrogen bonds with the substrate and ß-lactams, respectively. Our results provide valuable information on the molecular interactions essential for ligand recognition and catalysis by PBP6. Understanding these interactions will be helpful in the development of effective drugs to treat S. sonnei infections.

Development of Multiplex PCR Assay for Screening of T6SS-5 Gene Cluster: The Burkholderia pseudomallei Virulence Factor.

Despite the advanced understanding of the disease, melioidosis, an infection caused by Burkholderia pseudomallei, continues to be of global interest. The bacterial virulence factor, type six secretion system-5 (T6SS-5), in particular, is an essential factor for B. pseudomallei that is associated with internalization and intracellular survival of the pathogen. To detect the virulence gene cluster, this study has successfully developed a novel seven-gene (tssC-5, tagD-5, tssA-5, hcp-5, tssB-5, tssF-5, and vgrG-5) multiplex PCR assay. The optimum annealing temperature for this assay ranged between 59 and 62 °C. The limit of detection for this assay was 103 CFU/mL for all genes, excluding tssF-5, which was found at 105 CFU/mL of the bacterial concentration. In sensitivity and specificity tests, this multiplex assay was able to amplify all of the seven target genes from 93.8% (n = 33/35) clinical and 100% (n = 2/2) environmental isolates of B. pseudomallei. Whereas only four genes (tssC-5, tagD-5, tssF-5, and vgrG-5) were amplified from Bukholderia thailandesis, two genes (tagD-5 and tssB-5) were amplified from Bukholderia stagnalis, and zero target genes were amplified from Bukholderia ubonensis. No amplification of any genes was obtained when tested against isolated DNA from non-Bukholderia species (n = 20), which include Staphylococcus aureus, Klebsiella pneumoniae, Enterococcus faecalis, and others. In conclusion, this multiplex PCR assay is sensitive, species-specific, rapid, and reliable to detect the virulent gene cluster T6SS-5 of B. pseudomallei.