Rapid and accurate serotyping of Salmonella (S.) enterica subsp. enterica serovars is essential for effective public health surveillance, outbreak control, and food safety management. Traditional serotyping, although considered the gold standard, is time-consuming, technically demanding, and costly. This study aimed to evaluate the applicability of artificial intelligence (AI)-enhanced Fourier-transform infrared (FT-IR) spectroscopy using an IR Biotyper (Bruker Daltonics, Bremen, Germany) for the rapid and accurate serotyping of Salmonella enterica subsp. enterica isolates in Croatia. Materials and Methods: A total of 143 isolates representing different S. enterica serovars of human and food origin were analysed using the IR Biotyper. Each strain was tested in three biological and at least three technical replicates. The obtained results were compared with traditional serotyping according to the Kauffmann–White–Le Minor scheme. Isolate identification at the genus level was further confirmed by MALDI-TOF mass spectrometry. Results: The IR Biotyper demonstrated high reproducibility and complete concordance with standard serotyping methods, enabling accurate differentiation of the most prevalent S. enterica serovars in Croatia. Conclusions: Our findings demonstrate the applicability of FT-IR in routine laboratory work, with the potential to reduce typing time, reduce the number of strains, and lower overall costs required for epidemiological surveillance within the One Health approach.

Background/Objectives: Non-tuberculous mycobacteria (NTM) are increasingly recognized as important opportunistic pathogens at the human–animal–environment interface. Their growing relevance is driven by increasing disease burden, environmental persistence, occurrence in multiple animal hosts and complex antimicrobial resistance (AMR) patterns. Unlike classical zoonotic pathogens, most NTM are primarily acquired from shared environmental reservoirs rather than through sustained host-to-host transmission. This review examines NTM from a One Health perspective, focusing on AMR, ecology, animal occurrence, cross-species exposure and public health relevance. Methods: A narrative review of the current literature was conducted to synthesize evidence on the ecology, environmental reservoirs, occurrence in animals, transmission patterns and AMR mechanisms of NTM. Particular attention was given to studies addressing the human–animal–environment interface and the implications of NTM for One Health surveillance and risk assessment. Results: The reviewed literature shows that NTM are widely distributed in water, soil, sediments and biofilms, creating repeated opportunities for exposure in both animals and humans. They have been reported in livestock, wildlife, companion animals, reptiles and aquatic organisms, where they may act as colonizers, opportunistic pathogens, or sources of diagnostic interference. Evidence for direct animal-to-human transmission remains limited, but animal and environmental findings are important for understanding ecological overlap, host range and circulation of resistant strains. AMR in NTM is shaped by intrinsic resistance, acquired mutations, efflux activity, and biofilm-associated tolerance, which together complicate treatment and resistance prediction. Conclusions: NTM should be considered environmentally maintained, multi-host organisms of increasing One Health importance rather than conventional zoonotic pathogens. Improved interdisciplinary surveillance, diagnostics and research are needed to clarify exposure pathways, resistance development and public health risk.

Listeria monocytogenes is an important foodborne pathogen responsible for listeriosis, a severe human infection with high morbidity and mortality. Molecular serotyping is a universally accepted subtyping method for L. monocytogenes. Identification of strain serotype permits differentiation between significant foodborne strains (1/2a, 1/2b, and 4b) and allows a better understanding of their distribution and epidemiological significance. This study evaluated the implementation of real-time molecular serotyping of L. monocytogenes and compared its performance with conventional PCR methods. Eighty L. monocytogenes strains isolated from food and the food chain were analysed. Conventional multiplex PCR and qualitative real-time PCR (ReT-PCR) were used to identify molecular serogroups based on specific genetic markers to define the presence or absence of a particular serogroup of L. monocytogenes in strains. The results showed that ReT-PCR offers faster processing times than the conventional method. These strains belonged predominantly to serogroup IIa, followed by IIc, IIb, and IVb. ReT-PCR is becoming a standard tool for the detection and expression profiling of selected strains. This research highlights the importance of providing a rapid method for the qualitative detection of L. monocytogenes in food and integrating ReT-PCR into routine diagnostic applications.

This retrospective cross-sectional study investigated exposure to Leptospira spp. and Brucella canis in dogs housed in two animal shelters in Croatia. Archived serum samples from 375 dogs from the Dumovec shelter (Zagreb County) and the Friends Čakovec shelter (Međimurje County) were tested. Specific antibodies against Leptospira were detected by the microscopic agglutination test (MAT) using a panel of ten pathogenic serovars, while B. canis antibodies were screened using a rapid immunochromatographic assay. Associations with shelter, sex, age, serovar distribution, and antibody titers were statistically evaluated. Overall, 128/375 dogs (34.1%) were seropositive to at least one Leptospira serovar. Seropositivity was 29.6% in Dumovec and 36.1% in Čakovec, with no significant difference between shelters. No significant association was found between overall seropositivity and sex or age. Before adjustment for recent vaccination, the most frequent MAT reactions were directed against antigens representing Icterohaemorrhagiae, Australis, Canicola, Pomona, and Grippotyphosa. In a vaccination-related sensitivity analysis, MAT reactivity to antigens representing Australis, Pomona, and Grippotyphosa remained frequent, although this pattern should be interpreted cautiously because complete vaccine-product information was not consistently available. Most antibody titres were low to moderate, more consistent with previous exposure rather than with acute infection. No antibodies against B. canis were detected in any samples. These findings indicate substantial environmental exposure of shelter dogs in Croatia to pathogenic leptospires and support their role as sentinels of zoonotic risk. The predominance of MAT reactivity patterns associated with serogroups linked to wildlife and rodent reservoirs highlights the need for continued surveillance, rodent control, and careful interpretation of serology in relation to vaccination history.

Leptospirosis is a re-emerging zoonotic disease of increasing importance in both veterinary and public health, with its epidemiology strongly influenced by environmental and climatic factors. The aim of this study was to investigate the temporal and spatial distribution of canine leptospirosis in Croatia over an 11-year period (2013–2023), with particular emphasis on prevalence trends and serovar distribution. A total of 2,444 canine blood samples submitted for diagnostic testing were analysed using the microscopic agglutination test (MAT) for antibodies against 12 Leptospira serovars. Overall, 983 samples (40.22%) were seropositive. Annual prevalence showed marked fluctuations, with the highest numbers of positive samples recorded in 2018, 2020, and 2021. Geographical analysis revealed higher prevalence in central and northwestern Croatia, particularly in the City of Zagreb and surrounding counties. The most frequently detected serovars were Pomona (26.12%), Grippotyphosa (22.20%), Australis (16.96%), and Sejroe (14.95%), while other serovars occurred only sporadically. Temporal changes in serovar distribution were also observed, indicating dynamic epidemiological patterns. The identified fluctuations in prevalence are likely associated with environmental conditions favorable for Leptospira survival and transmission, including periods of increased precipitation and flooding documented in Croatia during the study period. These findings highlight considerable temporal, geographical, and serovar-related variability in canine leptospirosis, and emphasise the importance of continuous surveillance, regionally relevant vaccination strategies, and a One Health approach to disease prevention and control.

Contamination of ready‐to‐eat (RTE) foods with Listeria monocytogenes represents a significant public health concern, particularly due to the limited data available on its prevalence in traditional Croatian RTE products. This study is aimed at determining the prevalence of L. monocytogenes in traditional dried RTE meat products sampled from local markets in Zagreb, characterizing the genomic features of the isolated strains, and assessing their potential public health implications. A total of 50 RTE meat products, including Slavonian sausage, dry meat products, and garlic sausage, were collected and analyzed using standard cultural methods, followed by biochemical confirmation and serogroup identification with multiplex PCR. Whole‐genome sequencing (WGS) was employed to determine clonal complexes (CCs), sequence types (STs), and the presence of virulence‐associated genes. L. monocytogenes was detected in 22% (11/50) of the analyzed samples, of which 63.63% (7/11) were isolated from Slavonian sausage. Among the confirmed isolates, Serogroup IIa was the most prevalent (54.54%). WGS analysis revealed the presence of seven distinct CCs, of which CC9 ( n = 3), CC37 ( n = 2) and CC121 ( n = 2) were the most common. One of the strains was identified as CC1, which is considered hypervirulent. Single‐nucleotide polymorphism (SNP) analysis demonstrated that two strains identified as CC121 and two strains identified as CC9 were highly related (seven SNP differences in each). Overall, these findings provide new insights into the presence of L. monocytogenes in traditional RTE meat products in Zagreb and highlight a potential health risk for consumers.