PREVALENCE OF SALMONELLA IN CAPTIVE REPTILES FROM CROATIA.

Salmonellosis transmitted by pet reptiles is an increasing public health issue worldwide. The aim of this study was to investigate the prevalence of Salmonella strains from captive reptiles in Croatia. From November 2009 to November 2011 a total of 292 skin, pharyngeal, cloacal, and fecal samples from 200 apparently healthy reptiles were tested for Salmonella excretions by bacteriologic culture and serotyping. These 200 individual reptiles included 31 lizards, 79 chelonians, and 90 snakes belonging to private owners or housed at the Zagreb Zoo, Croatia. Salmonella was detected in a total of 13% of the animals, among them 48.4% lizards, 8.9% snakes, and 3.8% turtles. Representatives of five of the six Salmonella enterica subspecies were identified with the following proportions in the total number of isolates: Salmonella enterica enterica 34.6%, Salmonella enterica houtenae 23.1%, Salmonella enterica arizonae 23.1%, Salmonella enterica diarizonae 15.4%, and Salmonella enterica salamae 3.8%. The 14 different serovars isolated included several rarely occurring serovars such as Salmonella Apapa, Salmonella Halle, Salmonella Kisarawe, and Salmonella Potengi. These findings confirm that the prevalence of Salmonella is considerable in captive reptiles in Croatia, indicating that these animals may harbor serovars not commonly seen in veterinary or human microbiologic practice. This should be addressed in the prevention and diagnostics of human reptile-transmitted infections.

Characterization of egg laying hen and broiler fecal microbiota in poultry farms in Croatia, Czech Republic, Hungary and Slovenia.

Poultry meat is the most common protein source of animal origin for humans. However, intensive breeding of animals in confined spaces has led to poultry colonisation by microbiota with a zoonotic potential or encoding antibiotic resistances. In this study we were therefore interested in the prevalence of selected antibiotic resistance genes and microbiota composition in feces of egg laying hens and broilers originating from 4 different Central European countries determined by real-time PCR and 16S rRNA gene pyrosequencing, respectively. strA gene was present in 1 out of 10,000 bacteria. The prevalence of sul1, sul2 and tet(B) in poultry microbiota was approx. 6 times lower than that of the strA gene. tet(A) and cat were the least prevalent being present in around 3 out of 10,000,000 bacteria forming fecal microbiome. The core chicken fecal microbiota was formed by 26 different families. Rather unexpectedly, representatives of Desulfovibrionaceae and Campylobacteraceae, both capable of hydrogen utilisation in complex microbial communities, belonged among core microbiota families. Understanding the roles of individual population members in the total metabolism of the complex community may allow for interventions which might result in the replacement of Campylobacteraceae with Desulfovibrionaceae and a reduction of Campylobacter colonisation in broilers, carcasses, and consequently poultry meat products.

Microbes versus microbes: control of pathogens in the food chain.

Foodborne illness continues as a considerable threat to public health. Despite improved hygiene management systems and increased regulation, pathogenic bacteria still contaminate food, causing sporadic cases of illness and disease outbreaks worldwide. For many centuries, microbial antagonism has been used in food processing to improve food safety. An understanding of the mode of action of this microbial antagonism has been gained in recent years and potential applications in food and feed safety are now being explored. This review focuses on the potential opportunities presented, and the limitations, of using microbial antagonism as a biocontrol mechanism to reduce contamination along the food chain; including animal feed as its first link. © 2014 Society of Chemical Industry.

European validation of Real-Time PCR method for detection of Salmonella spp. in pork meat.

The classical microbiological method for detection of Salmonella spp. requires more than five days for final confirmation, and consequently there is a need for an alternative methodology for detection of this pathogen particularly in those food categories with a short shelf-life. This study presents an international (at European level) ISO 16140-based validation study of a non-proprietary Real-Time PCR-based method that can generate final results the day following sample analysis. It is based on an ISO compatible enrichment coupled to an easy and inexpensive DNA extraction and a consolidated Real-Time PCR assay. Thirteen laboratories from seven European Countries participated to this trial, and pork meat was selected as food model. The limit of detection observed was down to 10 CFU per 25 g of sample, showing excellent concordance and accordance values between samples and laboratories (100%). In addition, excellent values were obtained for relative accuracy, specificity and sensitivity (100%) when the results obtained for the Real-Time PCR-based methods were compared to those of the ISO 6579:2002 standard method. The results of this international trial demonstrate that the evaluated Real-Time PCR-based method represents an excellent alternative to the ISO standard. In fact, it shows an equal and solid performance as well as it reduces dramatically the extent of the analytical process, and can be easily implemented routinely by the Competent Authorities and Food Industry laboratories.

Findings of Devriesea agamarum associated infections in spiny-tailed lizards (Uromastyx sp.) in Croatia.

Actinobacteria are common agents that cause skin diseases in captive desert lizards, including the recently described Devriesea agamarum. To date, infections caused by D. agamarum, their symptoms, and treatment have been described only by the research group from Belgium that isolated the species in 2008. This article presents the symptoms that indicate the possibility of a D. agamarum-associated infection, such as scaly changes around the mouth in a juvenile lizard (Uromastyx ocelatta) and dermatitis in the form of skin scaling around the mouth and cloaca and over the dorsal part of the body in a group of four spiny-tailed lizards (Uromastyxgeyri). In two animals, swelling of the front limbs with the loss of some toes was also noted, a symptom not previously described with D. agamarum infections. Bacteriologic analysis of dermal lesion samples confirmed the presence of D. agamarum in all subjects. Treatment with ceftazidime was carried out, and the symptoms of dermatitis resolved, followed by negative bacteriologic findings. This is the first report, to our knowledge, that describes the diagnostics, detailed clinical picture with newly described symptoms, and treatment of lizards with D. agamarum-associated skin lesions that reside outside of Belgium. The results also confirm the effectiveness of the systemic administration of third-generation cephalosporin antibiotics in combination with local chlorhexidine in the treatment of D. agamarum infections.

Molecular characterization of atypical Chlamydia and evidence of their dissemination in different European and Asian chicken flocks by specific real-time PCR.

Chlamydia psittaci is a zoonotic pathogen associated primarily with avian chlamydiosis. New chlamydial agents with suspected zoonotic potential were recently detected from domestic poultry in Germany and France indicating that the spectrum of Chlamydiaceae encountered in birds is not confined to a single chlamydial species. For further characterization, a specific real-time PCR targeting the conserved 16S rRNA gene was developed and validated for a specific detection of these atypical Chlamydiaceae. In order to address the epidemiological importance of the new chlamydial agents and their distribution, Chlamydiaceae-positive chicken samples collected from flocks from five different countries were examined. The results confirmed that C.psittaci is not the predominant chlamydial species among chickens examined and suggested that the new chlamydial agents could putatively be widespread in poultry flocks (France, Greece, Croatia, Slovenia and China at least) justifying their systematic investigation when poultry samples are submitted to laboratories for avian chlamydiosis diagnosis. Besides, 16S rRNA-based dendrogram, including sequences from both isolates of the new chlamydial agents or positive samples as well as representative sequences from species belonging to the order Chlamydiales, showed the new chlamydial agents to form a distinct line of descent separated from those of other chlamydial species, but clearly grouped within the family Chlamydiaceae. Finally, the phylogenetic tree inferred from the multi-locus sequence typing based on four housekeeping fragments (gatA, gidA, enoA and hflX) and the ompA-based dendrogram showed an almost identical topology of the new chlamydial agents with that recovered by 16S rRNA-based dendrogram. Interestingly, partial ompA gene sequences displayed considerable diversity among isolates.

Genetic analysis of Newcastle disease virus strains isolated in Bosnia-Herzegovina, Croatia, Slovenia and Yugoslavia, reveals the presence of only a single genotype, V, between 1979 and 2002.

Newcastle disease (ND) epizootics in some European countries after the World War II were caused by ND virus (NDV) of multiple genotypes (IV-VIIa) occurring sequentially and/or simultaneously. This study was carried out to characterise the genetic composition of NDV strains during the outbreaks in the territory of the former Yugoslavia in order to enhance our understanding of the relationships of past epizootics in Europe. Sixty-eight NDV strains isolated between 1979 and 2002 were analysed by restriction enzyme digestion and partial sequencing of the fusion protein gene. All isolates were placed in genotype V, an exotic type, that was introduced to western Europe in 1970. Residue substitution analysis has allowed the recognition of four genetic variants, Vb1-Vb4, and the tracing of their movements. Vb1, a dominant variant in Bulgaria from the late 1970s, was also wide spread in the former Yugoslavia throughout the period under investigation. Vb2, a variant occurring in the neighbouring countries in the early 1970s could be the founder of the epidemic in Yugoslavia and it was present up to the late 1980s. Variants Vb3 and Vb4 could be found only after 1987. In conclusion, the ND outbreaks in Yugoslavia were part of the epizootic wave due to genotype V viruses that started in western Europe in 1970 and became endemic in the region. Inter-country transmission occurred for all variants, and Vb3 and Vb4 might have evolved during the endemic period.