Pathogens

Background: Numerous studies have reported on the epidemiology of hand, foot and mouth disease (HFMD) reinfection and its potential influencing factors; however, findings regarding reinfection rates as well as determinants such as gender, age, residence, and pathogens remain inconsistent. Due to this heterogeneity in reported outcomes, a comprehensive systematic review and meta-analysis are warranted to consolidate existing evidence. Methods: Effect estimates were expressed as reinfection rates, odds ratio (OR)/hazard ratio (HR) and 95% confidence intervals (CI). When necessary, data were converted to ensure consistency across comparison groups. Results: A thorough search was carried out using the predetermined literature retrieval approach across the PubMed, Web of Science, and Embase databases. Finally, 9 articles met the inclusion criteria and were included in this study. The results indicated that the overall reinfection rate for HFMD was 4.1% (95% CI: 2.0–6.2%). Males compared to females (overall effect = 1.256, 95% CI: 1.176–1.341), younger compared to older children (overall effect = 2.972, 95% CI: 1.512–5.843), scattered children compared to students (overall effect: 4.017, 95% CI: 1.560–10.344), and enterovirus 71 (EV71) compared to non-EV71 enteroviruses (overall effect = 0.71, 95% CI: 0.59–0.86) were associated with the HFMD reinfection. Conclusions: The overall HFMD reinfection rate was 4.1% (95% CI: 2.0–6.2%). Male, younger age, kindergarten children, and infection with non-EV71 enteroviruses (compared to EV71), were identified as significant risk factors for recurrent HFMD. Targeted intervention strategies should be developed for these high-risk populations to effectively reduce the incidence of reinfection.

L. monocytogenes is the causative agent of human listeriosis, a deadly disease with fatality rates up to 20%. L. monocytogenes has the ability to grow under harsh environmental conditions. It can form biofilms in food industries, making it capable of persisting in facilities. Given this scenario, it is of utmost importance to rapidly detect this bacterium not only in foods but also on food-contact surfaces. For the successful outcome of any given detection technology, it is imperative to properly process the samples. In the present work, PBS, LPT, and LPT-Pronase were compared to determine which one could provide better results in DNA-based detection. Additionally, the effect of a short TSB pre-enrichment was assessed. To better mimic a real scenario, L. monocytogenes monospecies and multispecies biofilms were analyzed. It was observed that supplementing LPT with pronase, a protein-degrading enzyme, could better detach the biofilm, which achieved a 0.5 cycle reduction compared to the other broths, and the pre-enrichment reduced the real-time PCR by ~2 cycles. The samples were analyzed by real-time PCR and colorimetric LAMP, and the same results were obtained with both techniques regardless of the concentration of L. monocytogenes present in the biofilm; the initial concentration was 1.8 log CFU/cm² 15 min after the pre-enrichment. The results were confirmed by real-time PCR, which demonstrated the applicability of the methodology to be applied in decentralized setups, such as food-processing facilities, with minimal laboratory infrastructure.

Background: Enterovirus-D68 (EV-D68) was long underreported, with only sporadic cases of respiratory disease worldwide until 2014, when numerous countries experienced significant outbreaks of EV-D68. In Croatia, sporadic detections have primarily resulted from an absence of systematic surveillance. Following the increased incidence of EV-D68 across Europe in 2022, we started to characterize EV-positive respiratory samples in Zagreb to confirm the presence of EV-D68 and identify circulating lineages through phylogenetic analysis. Methods: Respiratory samples from individuals with acute respiratory infection and additional clinical symptoms were tested at the Virology Laboratory of the Croatian Institute of Public Health, and EV-positive samples were further screened using the real-time RT-qPCR method for EV-D68. VP1 sequences were obtained by sequencing and subsequently genotyped. Results: Between March 2022 and December 2024, EV was detected in 2048 respiratory samples. Annual distributions of EV detections were 656 (10.0%) in 2022, 785 (8.1%) in 2023, and 607 (7.4%) in 2024. EV-D68 was identified in 13.1% of EV-positive samples in 2022, 1.4% in 2023, and 19.6% in 2024. The peaks in EV-D68 circulation were observed in July (n = 24) and September (n = 24) in 2022 and in September 2024 (n = 62). Phylogenetic analysis of EV-D68 VP1 sequences revealed the presence of two major clades, A2 and B3. The sequences from 2022 clustered exclusively within clade B3, while in 2024 A2 clade was newly introduced. Conclusions: We confirmed the presence of EV-D68 in Croatia with circulating lineages corresponding to those detected elsewhere in Europe. The absence of routine testing has likely led to an underestimation of EV-D68 prevalence. These findings underscore the urgent need for ongoing surveillance and genomic characterization to clarify EV-D68 epidemiology in Croatia.