Purpose: This prospective, longitudinal study aims to evaluate the durability and functionality of SARS-CoV-2 Ancestral strain (Wuhan-Hu-1)-specific immune responses induced by COVID-19 vaccination and natural infection over a 12-month period. This article reviews the study protocol, design, methodology, ongoing data collection, analysis procedures, and demographic characteristics of the cohort enrolled. Participants: Between March 2021 and May 2022, 400 participants were enrolled with a 12-month follow-up, concluding in May 2023. Two main groups of participants: (1) serologically SARS-CoV-2-naïve individuals receiving the BNT162b2 primary series vaccination (referred to as VAC) and (2) those who recently recovered from COVID-19 infection within 30 days, regardless of vaccination history (referred to as COV). Additionally, a subset of 45 participants with selected COVID-19 exposure histories provided peripheral blood mononuclear cells (PBMCs) for cross-sectional analysis six months after enrollment. Findings to date: Out of 400 participants, 66.8% (n=267) completed the follow-up. Among them, 52.8% (n=141) were in VAC, and 47.2% (n=126) were in COV. As the study progressed, we acknowledged cross-over between initial groups, leading to restructuring into five revised groups based on sequential exposure events. Sociodemographic factors revealed statistically significant age distribution differences (p=0.001) in both initial and revised groups, with no significant differences observed for sex. Future plans: LONGTONG-SARS2 assesses the host-pathogen interactions central to the development of COVID-19 immunity. With enrollment spanning two years of the pandemic, most participants exhibited mixed SARS-CoV-2 exposures—via vaccination and infection—resulting in diverse subgroups of interest. Notably, the inclusion of SARS-CoV-2-naïve, pre-exposure serum samples allowed for robust comparator and reduced potential biases. Ongoing analyses will include serology kinetics, memory cells ELISpots, B cells repertoire analysis, cytokine/chemokine profiling, and proteomic pathway to comprehensively examine the immune response against the SARS-CoV-2, thus informing and potentially predicting dynamic longitudinal responses against new more transmissible, immune-evasive SARS-CoV-2 variants. STRENGTH AND LIMITATIONS OF THIS STUDY: - LONGTONG-SARS2 is a prospective longitudinal study that comprehensively evaluates the SARS-CoV-2 immune response among a diverse group of individuals, stratified based on the sequential order of SARS-CoV-2 exposure events, whether from COVID-19 vaccination or infection. - Pre-vaccination serum samples were collected from serologically SARS-CoV-2 naive individuals scheduled to receive the BNT162b2 primary series vaccination during the initial mass COVID-19 vaccination phase in Malaysia in early 2021. - The longitudinal serum sample collection spanned two years of the COVID-19 pandemic, from March 2021 to May 2023. This extended duration allows for robust monitoring of the immune response against SARS-CoV-2 variants in comparison to the ancestral strain. - There is a risk of misclassification of some individuals’ SARS-CoV-2 exposure status through serology, as certain sampling timepoints had intervals of three months. Additionally, our study relies on self-reported data through the MySejahtera application (Malaysia’s electronic medical record by the Ministry of Health) for second confirmation, potentially leading to underdiagnosed and underreported cases of asymptomatic infection.

Urban stray cats are often associated with ectoparasites and zoonotic pathogens due to their unsanitary living conditions and lack of veterinary care. Fleas, especially Ctenocephalides spp., are competent vectors of Bartonella spp., a genus of emerging bacterial pathogens with both public health and veterinary relevance. This study investigated the presence of Bartonella DNA in ectoparasitic fleas infesting stray cats in various urban habitats within the Klang Valley, Malaysia. A total of 204 fleas were collected from 89 stray cats. Fleas were identified morphologically using established taxonomic keys under a light microscope and further validated through PCR amplification of the mitochondrial cytochrome c oxidase subunit I (cox1) gene. Detection of Bartonella spp. was conducted by targeting the citrate synthase (gltA) gene. All fleas were confirmed as Ctenocephalides felis, with an infestation prevalence of 39.33% among the cats sampled. Of 118 C. felis specimens tested, 86.44% were positive for Bartonella DNA, one of the highest worldwide and significantly surpassing previous Malaysian reports. Sequencing of 12 positive samples showed identities with B. claridgeiae (58.3%), B. henselae (25.0%), an uncultured Bartonella species (8.3%) and a Bartonella isolate from a dog in Chile (8.3%). These results highlight the significant presence of Bartonella, causative agent of cat-scratch disease in stray cats, emphasizing their potential role as urban reservoirs and vectors. The findings underscore the need for ectoparasite surveillance and zoonotic pathogen control as integral components of stray animal management in Malaysia’s urban settings.

Understanding how clinical symptoms relate to immune responses during major variant transitions remains important for informing post-pandemic surveillance and vaccination strategies. This study compared symptom patterns and SARS-CoV-2-specific anti-RBD IgM and anti-S1 IgG antibody responses among vaccinated individuals infected during the pre-Omicron and Omicron-dominant periods, representing a key phase in the evolution of SARS-CoV-2 population immunity. A retrospective analysis of 216 confirmed COVID-19 cases was performed by evaluating 11 predefined symptoms together with anti-RBD IgM and anti-S1 IgG levels measured at Day-14 after symptom onset, corresponding to the period when humoral antibody responses are detectable following SARS-CoV-2 infection. Participants with breakthrough infection during the Omicron-dominant period reported fewer symptoms overall compared to the pre-Omicron period, with a median of three versus four symptoms, respectively. Cough was the most common symptom during the Omicron period (82.1%), followed by sore throat (81.4%) and fever (78.6%). In contrast, loss of taste or smell was significantly more frequent in the pre-Omicron period (64.8% versus 22.9%, p < 0.05). IgG levels were significantly higher during the Omicron period than during the pre-Omicron period, increasing by 42.3%, reflecting enhanced antibody responses likely driven by repeated exposure. A consistent association between cough and elevated IgG levels was observed in both periods (p < 0.05), suggesting an association between symptom presentation and the magnitude of the early humoral response. These findings suggest that while clinical symptom profiles evolved across a major SARS-CoV-2 variant transition, certain symptom–antibody relationships remained consistent. Such associations may provide insight into how clinical manifestations relate to immune responses in populations with pre-existing immunity and may support interpretation of symptomatic infection during ongoing SARS-CoV-2 circulation.