The immune response to a Coxiella burnetii vaccine in sheep varies according to their natural pre-exposure.

Q fever in humans is caused by Coxiella (C.) burnetii. In 2008 and 2012, cases of Q fever in humans were linked to an infected flock of approximately 650 ewes. Since 2013 gimmers (G'13, G'14, G'15 etc.) were primary vaccinated (two doses) with an inactivated C.burnetii vaccine without any revaccination. In 2013, 30 ewes were primary vaccinated (A'13). Shedding was annually monitored by qPCR-testing of vaginal and nasal swabs collected at lambing. Animals were tested for Phase I- (PhI) and PhII-antibodies (Ab) and for PhII-specific-interferon-γ (IFN-γ) before and after vaccination. The effect of a revaccination was determined in 2018 and 2023. Groups of randomly selected gimmers primary vaccinated in 2015, 2016 and 2017 and a mixed group of older animals (A'13, G'13 and G'14) were revaccinated once in 2018. The trial was repeated in 2023 on groups primary vaccinated in 2019-2023. Major shedding after the outbreak in 2012 ceased in 2014. Thereafter C.burnetii was only sporadically detected at low-level in 2018, 2021 and 2023. Sheep naturally exposed to C.burnetii during the outbreak in 2012 (A'13, G'13) mounted a strong and complete (PhI, PhII, IFN-γ) recall immune response after vaccination. A serological PhI+/PhII+ pattern dominated after vaccination. In contrast, since 2014 a weaker immune response (PhII-titre, IFN-γ) and a dominance of the PhI-/PhII+ pattern was observed in vaccinated gimmers. The number of serologically non-responding gimmers to vaccination increased to 25.0 % in G'16/G'17 and 40.4 % in G'19/G'20. But revaccination even three (G'15 in 2018) and four (G'19 in 2023) years after primary vaccination resulted in a strong and complete immune response. No difference of the immune response nor to more recently primary vaccinated animals (G'23 in 2023) nor to those animals that were present during the outbreak (A'13/G'13/G'14 in 2018) was observed.

Seroprevalence of Brucella Infection in Wild Boars (Sus scrofa) of Bavaria, Germany, 2019 to 2021 and Associated Genome Analysis of Five B. suis Biovar 2 Isolates.

Brucella species are highly pathogenic zoonotic agents and are found in vertebrates all over the world. To date, Germany is officially declared free from brucellosis and continuous surveillance is currently limited to farm ruminants. However, porcine brucellosis, mostly caused by B. suis biovar 2, is still found in wild boars and hares. In the present study, a three-year monitoring program was conducted focusing on the wild boar population in the state of Bavaria. Serologic screening of 11,956 animals and a direct pathogen detection approach, including a subset of 681 tissue samples, was carried out. The serologic incidence was 17.9%, which is in approximate accordance with previously published results from various European countries. Applying comparative whole genome analysis, five isolated B. suis biovar 2 strains from Bavaria could be assigned to three known European genetic lineages. One isolate was closely related to another strain recovered in Germany in 2006. Concluding, porcine brucellosis is endemic in Bavaria and the wild boar population represents a reservoir for genetically distinct B. suis biovar 2 strains. However, the transmission risk of swine brucellosis to humans and farm animals is still regarded as minor due to low zoonotic potential, awareness, and biosafety measures.

CD31, EDNRB and TSPAN7 are promising prognostic markers in clear-cell renal cell carcinoma revealed by genome-wide expression analyses of primary tumors and metastases.

Currently used clinicopathological parameters are insufficient for a reliable prediction of metastatic risk and disease-free survival (DFS) of patients with clear-cell renal cell carcinoma (ccRCC). To identify prognostic genes, the expression profiles of primary ccRCC obtained from patients with different DFS--eight synchronously, nine metachronously and seven not metastasized tumors--were determined by genome-wide expression analyses. Synchronously and metachronously metastasized primary ccRCC differed in the expression of 167 genes. Thirty-six of these genes were also differentially expressed in synchronously vs. metachronously developed pulmonary metastases analyzed in a previous study. Because of their DFS-associated deregulation that is concordant in metastases and primary ccRCC, these genes are potentially functionally involved in metastatic tumor growth and are also prognostically useful. A prognostic impact was confirmed for the genes CD31, EDNRB and TSPAN7 at the mRNA level (n=86), and for TSPAN7 at the protein level (n=106). Patients with a higher gene expression of EDNRB or TSPAN7, or with TSPAN7-positive vessels in both cores investigated on tissue microarrays had a significantly longer DFS and tumor-specific survival (TSS). Patients with a higher CD31 gene expression showed a significantly longer TSS. EDNRB was an independent prognostic marker for the DFS. CD31, EDNRB and TSPAN7 had an independent impact on the TSS. In summary, comparative analysis of primary tumors and metastases is appropriate to identify independent prognostic markers in ccRCC. Gene expression of CD31 and EDNRB, and endothelial TSPAN7 protein level are potentially useful to improve outcome prediction because of their independent prognostic impact.

Long-term control of Coxiellosis in sheep by annual primary vaccination of gimmers.

Coxiella (C.) burnetii, a Gram-negative intracellular bacterium, causes Q fever in humans and Coxiellosis in animals. Ruminants are a primary source of human infection with C.burnetii. In 2013, vaccination was implemented in a sheep flock with 650 ewes associated with two outbreaks of Q fever in humans in 2008 and 2012. Only gimmers (yearlings) received two doses of a commercial C.burnetii phase I whole cell vaccine three weeks apart (primary vaccination) without any revaccination. Vaginal and nasal swabs collected shortly after lambing were tested by qPCR. Additionally, a group of non-vaccinated sentinels was serologically monitored for phase I (PhI), II (PhII) antibodies and for Interferon γ (IFN-γ) after stimulation of whole blood cells with PhII-antigen with and without an IL-10-neutralizing monoclonal antibody. In 2021, 679 sera collected in 2014-2021 were retested retrospectively with three commercial ELISA kits and one batch of an in-house PhI/PhII-ELISA. A low-level shedding of C.burnetii (<103 mean C.burnetii/swab) was observed until 2014. In 2021 C.burnetii was detected in two animals (<103.1C.burnetii/swab), but vaginal swabs collected at two subsequent lambing seasons remained negative. Seroconversion of sentinels was detected until 2017. However, the retrospective analysis of sentinels in 2021 revealed additional single seropositive animals from 2018 to 2021. IFN-γ reactivity was observed during the whole study period; it peaked in 2014 and in 2018 and decreased thereafter. The sporadic detection of C.burnetii and the immune responses of sentinels suggested that a subliminal infection persisted despite vaccination. Nevertheless, vaccination of gimmers prevented the development of a major outbreak, it controlled the infection and reduced the risk of human infection.

Antimicrobial Resistance, Serologic and Molecular Characterization of E. coli Isolated from Calves with Severe or Fatal Enteritis in Bavaria, Germany.

Worldwide, enterotoxigenic Escherichia coli (ETEC) cause neonatal diarrhea and high mortality rates in newborn calves, leading to great economic losses. In Bavaria, Germany, no recent facts are available regarding the prevalence of virulence factors or antimicrobial resistance of ETEC in calves. Antimicrobial susceptibility of 8713 E. coli isolates obtained from 7358 samples of diseased or deceased diarrheic calves were investigated between 2015 to 2019. Considerably high rates of 84.2% multidrug-resistant and 15.8% extensively drug-resistant isolates were detected. The resistance situation of the first, second and third line antimicrobials for the treatment, here amoxicillin-clavulanate, enrofloxacin and trimethoprim-sulfamethoxazole, is currently acceptable with mean non-susceptibility rates of 28.1%, 37.9% and 50.0% over the investigated 5-year period. Furthermore, the ETEC serotypes O101:K28, O9:K35, O101:K30, O101:K32, O78:K80, O139:K82, O8:K87, O141:K85 and O147:K89, as well as the virulence factors F17, F41, F5, ST-I and stx1 were identified in a subset of samples collected in 2019 and 2020. The substantially high rates of multi- and extensively drug-resistant isolates underline the necessity of continuous monitoring regarding antimicrobial resistance to provide reliable prognoses and adjust recommendations for the treatment of bacterial infections in animals.

ATP-Dependent C-F Bond Cleavage Allows the Complete Degradation of 4-Fluoroaromatics without Oxygen.

UNLABELLED: Complete biodegradation of the abundant and persistent fluoroaromatics requires enzymatic cleavage of an arylic C-F bond, probably the most stable single bond of a biodegradable organic molecule. While in aerobic microorganisms defluorination of fluoroaromatics is initiated by oxygenases, arylic C-F bond cleavage has never been observed in the absence of oxygen. Here, an oxygen-independent enzymatic aryl fluoride bond cleavage is described during the complete degradation of 4-fluorobenzoate or 4-fluorotoluene to CO2 and HF in the denitrifying Thauera aromatica: the ATP-dependent defluorination of 4-fluorobenzoyl-coenzyme A (4-F-BzCoA) to benzoyl-coenzyme A (BzCoA) and HF, catalyzed by class I BzCoA reductase (BCR). Adaptation to growth with the fluoroaromatics was accomplished by the downregulation of a promiscuous benzoate-CoA ligase and the concomitant upregulation of 4-F-BzCoA-defluorinating/dearomatizing BCR on the transcriptional level. We propose an unprecedented mechanism for reductive arylic C-F bond cleavage via a Birch reduction-like mechanism resulting in a formal nucleophilic aromatic substitution. In the proposed anionic 4-fluorodienoyl-CoA transition state, fluoride elimination to BzCoA is favored over protonation to a fluorinated cyclic dienoyl-CoA. IMPORTANCE: Organofluorides are produced as pesticides, pharmaceuticals, and other chemicals and comprise approximately one quarter of all organic compounds in the pharmaceutical and agricultural sectors; they are considered a growing class of environmentally relevant persistent pollutants. Especially in the case of fluoroaromatics, biodegradation is hampered by the extreme stability of the arylic C-F bond. In aerobic microorganisms, degradation proceeds via oxygenase-dependent C-F bond cleavage reactions, whereas the enzymes involved in the degradation of fluoroaromatics at anoxic sites are unknown. Here we report a strategy for the complete biodegradation of a fluoroaromatic to CO2 and HF in a denitrifying bacterium via activation to a CoA ester, followed by oxygen-independent arylic C-F bond cleavage catalyzed by an ATP-dependent enzyme. This reaction, in conjunction with a transcriptional adaptation to fluorinated growth substrates, is essential for the anoxic biodegradation of 4-fluorobenzoate/4-F-toluene and probably other fluoroaromatics.

Bioinformatics for RNomics.

Rapid improvements in high-throughput experimental technologies make it nowadays possible to study the expression, as well as changes in expression, of whole transcriptomes under different environmental conditions in a detailed view. We describe current approaches to identify genome-wide functional RNA transcripts (experimentally as well as computationally), and focus on computational methods that may be utilized to disclose their function. While genome databases offer a wealth of information about known and putative functions for protein-coding genes, functional information for novel non-coding RNA genes is almost nonexistent. This is mainly explained by the lack of established software tools to efficiently reveal the function and evolutionary origin of non-coding RNA genes. Here, we describe in detail computational approaches one may follow to annotate and classify an RNA transcript.