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Beta-lactamase resistant bacteria and especially ESBL producing Enterobacteriaceae are an increasing problem worldwide. For this reason a major interest in efficient and reliable methods for rapid screening of high sample numbers is recognizable. Therefore, a multiplex real-time PCR was developed to detect the predominant class A beta-lactamase genes blaCTX-M, blaSHV, blaTEM and CIT-type AmpCs in a one-step reaction. A set of 114 Enterobacteriaceae containing previously identified resistance gene subtypes and in addition 20 undefined animal and environmental isolates were used for the validation of this assay. To confirm the accessibility in variable settings, the real-time runs were performed analogous in two different laboratories using different real-time cyclers. The obtained results showed complete accordance between the real-time data and the predetermined genotypes. Even if sequence analyses are further necessary for a comprehensive characterization, this method was proofed to be reliable for rapid screening of high sample numbers and therefore could be an important tool for e. g. epidemiological purposes or support infection control measures.

Since the first description of a plasmid-mediated colistin resistance gene (mcr-1) in November 2015 multiple reports of mcr-1 positive isolates indicate a worldwide spread of this newly discovered resistance gene in Enterobacteriaceae. Although the occurrence of mcr-1 positive isolates of livestock, food, environment and human origin is well documented only few systematic studies on the prevalence of mcr-1 are available yet. Here, comprehensive data on the prevalence of mcr-1 in German livestock and food isolates are presented. Over 10.600 E. coli isolates from the national monitoring on zoonotic agents from the years 2010-2015 were screened for phenotypic colistin resistance (MIC value >2 mg/l). Of those, 505 resistant isolates were screened with a newly developed TaqMan-based real-time PCR for the presence of the mcr-1 gene. In total 402 isolates (79.8% of colistin resistant isolates) harboured the mcr-1 gene. The prevalence was depending on the food production chain. The highest prevalence was detected in the turkey food chain (10.7%), followed by broilers (5.6%). A low prevalence was determined in pigs, veal calves and laying hens. The mcr-1 was not detected in beef cattle, beef and dairy products in all years investigated. In conclusion, TaqMan based real-time PCR provides a fast and accurate tool for detection of mcr-1 gene. The overall detection rate of 3.8% for mcr-1 among all E. coli isolates tested is due to high prevalence of mcr-1 in poultry production chains. More epidemiological studies of other European countries are urgently needed to assess German prevalence data.

Enveloped respiratory viruses, including the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), can be transmitted through aerosols and contact with contaminated surfaces. The stability of these viruses outside the host significantly impacts their transmission dynamics and the spread of diseases. In this study, we investigated the tenacity of Feline Coronavirus (FCoV) in aerosols and on surfaces under varying environmental conditions. We found that airborne FCoV showed different stability depending on relative humidity (RH), with higher stability observed at low and high RH. Medium RH conditions (50–60%) were associated with increased loss of infectivity. Furthermore, FCoV remained infectious in the airborne state over 7 h. On stainless-steel surfaces, FCoV remained infectious for several months, with stability influenced by organic material and temperature. The presence of yeast extract and a temperature of 4 °C resulted in the longest maintenance of infectivity, with a 5 log 10 reduction of the initial concentration after 167 days. At 20 °C, this reduction was achieved after 19 days. These findings highlight the potential risk of aerosol and contact transmission of respiratory viruses, especially in enclosed environments, over extended periods. Studying surrogate viruses like FCoV provides important insights into the behavior of zoonotic viruses like SARS-CoV-2 in the environment.

Antimicrobial multidrug-resistant microorganisms (MDRO) can be transmitted between companion animals and their human owners. Aim of this study was to determine the prevalence of extended spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-E) and Staphylococcus aureus including methicillin-resistant S. aureus (MRSA) in different companion animal species. Dogs (n = 192), cats (n = 74), and rabbits (n = 17), treated in a veterinary practice and hospital or living in an animal shelter and private households, were sampled. All facilities were located in a region characterized by a high density of pig production. Nasal, buccal and perianal swabs were enriched and cultured on solid chromogenic selective media. A subgroup of 20 animals (13 dogs, 3 cats, 4 rabbits) was analyzed for the presence of staphylococci other than S. aureus. Amongst all animals (n = 283), twenty dogs (10.4%) and six cats (8.1%) carried S. aureus. MRSA was found in five dogs (2.6%) and two cats (2.7%). Isolates were of spa types t011, t034, t108 (all mecA-positive, ST398), and t843 (mecC-positive, ST130), typical for livestock-associated (LA)-MRSA. Except for one dog, MRSA-positive animals did not have direct contact to husbandry. ESBL-Escherichia coli (blaCTX-M/blaTEM/blaSHV genes) were present in seven dogs (3.6%), one cat (1.4%) possessed a cefotaxime-resistant Citrobacter freundii isolate (blaTEM/blaCMY-2 genes). MDRO carriage was associated with animals from veterinary medical settings (p<0.05). One dog and one rabbit carried methicillin-resistant coagulase-negative staphylococci. The exclusive occurrence of MRSA lineages typically described for livestock stresses the impact of MDRO strain dissemination across species barriers in regional settings. Presence of ESBL-E and LA-MRSA among pets and probable dissemination in clinical settings support the necessity of a \"One Health\" approach to address the potential threats due to MDRO-carrying companion animals.

Antimicrobial-resistant bacteria originating from broilers pose an ongoing challenge as they can spread in the environment and food chain. One approach to lower colonization is to administer live bacteria to broilers. A live Escherichia coli (E. coli) vaccine, consisting of a single E. coli strain used as a competitive exclusion (CE) approach, was evaluated to decrease the colonization with ESBL-/pAmpC- producing E. coli . 168 ESBL-/pAmpC- negative Ranger Gold broilers were divided into six groups (3 x n  = 46 experimental groups; 3 x n  = 10 control groups for quality assurance). The E. coli vaccine was administered on day one through coarse spray or by drinking water on day five. Experimental groups were orally co-colonized with 10 2 cfu of one ESBL- (ST410, bla CTX−M−15 ) and one pAmpC- producing E. coli (ST10, bla CMY−2 / mcr -1) on day three. Colonization status was monitored throughout the trial and quantified at the end of the study (day 49). A transient reduction in ESBL-producing E. coli colonization ( p  < 0.001) was observed following coarse spray administration. However, this decrease was not sustained over time. It can be concluded that a single E. coli strain originating from a live vaccine cannot decrease colonization of broilers with ESBL-/pAmpC-  E. coli throughout a fattening period.

Invertebrate pests and parasites of humans, animals, and plants continue to cause serious diseases and remain as a high treat to agricultural productivity and storage. The rapid and accurate species identification of the pests and parasites are needed for understanding epidemiology, monitoring outbreaks, and designing control measures. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) profiling has emerged as a rapid, cost effective, and high throughput technique of microbial species identification in modern diagnostic laboratories. The development of soft ionization techniques and the release of commercial pattern matching software platforms has resulted in the exponential growth of applications in higher organisms including parasitology. The present review discusses the proof-of-principle experiments and various methods of MALDI MS profiling in rapid species identification of both laboratory and field isolates of pests, parasites and vectors.

The awareness of non-malarial febrile illnesses (NMFIs) has been on the rise over the last decades. Therefore, we undertook a systematic literature review and meta-analysis of causative agents of non-malarial fevers on the African continent. We searched for literature in African Journals Online, EMBASE, PubMed, Scopus, and Web of Science databases to identify aetiologic agents that had been reported and to determine summary estimates of the proportional morbidity rates (PMr) associated with these pathogens among fever patients. A total of 133 studies comprising 391,835 patients from 25 of the 54 African countries were eligible. A wide array of aetiologic agents were described with considerable regional differences among the leading agents. Overall, bacterial pathogens tested from blood samples accounted for the largest proportion. The summary estimates from the meta-analysis were low for most of the agents. This may have resulted from a true low prevalence of the agents, the failure to test for many agents or the low sensitivity of the diagnostic methods applied. Our meta-regression analysis of study and population variables showed that diagnostic methods determined the PMr estimates of typhoidal Salmonella and Dengue virus. An increase in the PMr of Klebsiella spp. infections was observed over time. Furthermore, the status of patients as either inpatient or outpatient predicted the PMr of Haemophilus spp. infections. The small number of epidemiological studies and the variety of NMFI agents on the African continent emphasizes the need for harmonized studies with larger sample sizes. In particular, diagnostic procedures for NMFIs should be standardized to facilitate comparability of study results and to improve future meta-analyses. Reliable NMFI burden estimates will inform regional public health strategies.

The colonization of broilers with extended-spectrum β-lactamase- (ESBL-) and plasmid-mediated AmpC β-lactamase- (pAmpC-) producing Enterobacteriaceae has been extensively studied. However, only limited data on intervention strategies to reduce the colonization throughout the fattening period are available. To investigate practically relevant management measures for their potential to reduce colonization, a recently published seeder-bird colonization model was used. Groups of 90 broilers (breed Ross 308) were housed in pens under conventional conditions (stocking of 39 kg/m 2 , no enrichment, water and feed ad libitum ). Tested measures were investigated in separate trials and included (I) an increased amount of litter in the pen, (II) the reduction of stocking density to 25 kg/m 2 , and (III) the use of an alternative broiler breed (Rowan x Ranger). One-fifth of ESBL- and pAmpC- negative broilers (n = 18) per group were orally co-inoculated with two E . coli strains on the third day of the trial (seeder). One CTX-M-15-positive E . coli strain (ST410) and one CMY-2 and mcr-1-positive E . coli strain (ST10) were simultaneously administered in a dosage of 10 2 cfu. Colonization of all seeders and 28 non-inoculated broilers (sentinel) was assessed via cloacal swabs during the trials and a final necropsy at a target weight of two kilograms (= d 36 (control, I-II), d 47 (III)). None of the applied intervention measures reduced the colonization of the broilers with both the ESBL- and the pAmpC- producing E . coli strains. A strain-dependent reduction of colonization for the ESBL- producing E . coli strain of ST410 by 2 log units was apparent by the reduction of stocking density to 25 kg/m 2 . Consequently, the tested management measures had a negligible effect on the ESBL- and pAmpC- colonization of broilers. Therefore, intervention strategies should focus on the prevention of ESBL- and pAmpC- colonization, rather than an attempt to reduce an already existing colonization.

Background Broiler chickens are frequently colonized with Extended-Spectrum Beta-Lactamase- (ESBL-) and plasmid mediated AmpC Beta-Lactamase- (pAmpC-) producing Enterobacterales, and we are confronted with the potential spread of these resistant bacteria in the food chain, in the environment, and to humans. Research focused on identifying of transmission routes and investigating potential intervention measures against ESBL- and pAmpC- producing bacteria in the broiler production chain. However, few data are available on the effects of cleaning and disinfection (C&D) procedures in broiler stables on ESBL- and pAmpC- producing bacteria. Results We systematically investigated five broiler stables before and after C&D and identified potential ESBL- and pAmpC- colonization sites after C&D in the broiler stables, including the anteroom and the nearby surrounding environment of the broiler stables. Phenotypically resistant E. coli isolates grown on MacConkey agar with cefotaxime were further analyzed for their beta-lactam resistance genes and phylogenetic groups, as well as the relation of isolates from the investigated stables before and after C&D by whole genome sequencing. Survival of ESBL- and pAmpC- producing E. coli is highly likely at sites where C&D was not performed or where insufficient cleaning was performed prior to disinfection. For the first time, we showed highly related ESBL-/pAmpC- producing E. coli isolates detected before and after C&D in four of five broiler stables examined with cgMLST. Survival of resistant isolates in investigated broiler stables as well as transmission of resistant isolates from broiler stables to the anteroom and surrounding environment and between broiler farms was shown. In addition, enterococci (frequently utilized to detect fecal contamination and for C&D control) can be used as an indicator bacterium for the detection of ESBL-/pAmpC- E. coli after C&D. Conclusion We conclude that C&D can reduce ESBL-/pAmpC- producing E. coli in conventional broiler stables, but complete ESBL- and pAmpC- elimination does not seem to be possible in practice as several factors influence the C&D outcome (e.g. broiler stable condition, ESBL-/pAmpC- status prior to C&D, C&D procedures used, and biosecurity measures on the farm). A multifactorial approach, combining various hygiene- and management measures, is needed to reduce ESBL-/pAmpC- E. coli in broiler farms.