Low temperature preservation of porcine semen: influence of short antimicrobial lipopeptides on sperm quality and bacterial load.

Antimicrobial resistance is a steadily increasing problem and poses a serious threat to global public health. Therefore, it is highly necessary to advance the development of novel antimicrobial compounds and semen preservation strategies. The aim of this study was to evaluate a low temperature, antibiotic-free preservation procedure using Androstar Premium (ASP) extender (Minitüb) with antimicrobial lipopeptides. Firstly, seven lipopeptides in two concentrations (1 × minimum inhibitory concentration (MIC)/2 × MIC) were tested on their sperm-compatibility at 17 °C. Two lipopeptides, C16-KKK-NH2 and C16-KKKK-NH2, did not negatively affect sperm quality and were further evaluated for their efficiency of bacterial growth inhibition at 5 °C. Besides an overall diminution of colony forming units, both peptides showed a reduction of bacterial subcultures (n = 103) with a decrement in Gram-positive rods from 65 (ASP w/o supplements) to 39/52 (ASP w/ C16-KKK-NH2/C16-KKKK-NH2), in Gram-positive cocci from 21 to 9/10 and in Gram-negative species from 17 to 8/5 total subcultures. Furthermore, lipopeptides revealed activity towards selected bacteria of potential concern in artificial insemination like Trueperella pyogenes, Alcaligenes faecalis, Pseudomonas aeruginosa (not C16-KKK-NH2), Pasteurella sp., Providencia stuartii, Escherichia coli (not C16-KKKK-NH2) and Streptococcus porcinus (not C16-KKKK-NH2). Consequently, both tested lipopeptides are promising candidates for alternative antibiotic-free preservation techniques of boar semen.

Factors determining microbial colonization of liquid nitrogen storage tanks used for archiving biological samples.

The availability of bioresources is a precondition for life science research, medical applications, and diagnostics, but requires a dedicated quality management to guarantee reliable and safe storage. Anecdotal reports of bacterial isolates and sample contamination indicate that organisms may persist in liquid nitrogen (LN) storage tanks. To evaluate the safety status of cryocollections, we systematically screened organisms in the LN phase and in ice layers covering inner surfaces of storage tanks maintained in different biobanking facilities. We applied a culture-independent approach combining cell detection by epifluorescence microscopy with the amplification of group-specific marker genes and high-throughput sequencing of bacterial ribosomal genes. In the LN phase, neither cells nor bacterial 16S rRNA gene copy numbers were detectable (detection limit, 102 cells per ml, 103 gene copies per ml). In several cases, small numbers of bacteria of up to 104 cells per ml and up to 106 gene copies per ml, as well as Mycoplasma, or fungi were detected in the ice phase formed underneath the lids or accumulated at the bottom. The bacteria most likely originated from the stored materials themselves (Elizabethingia, Janthibacterium), the technical environment (Pseudomonas, Acinetobacter, Methylobacterium), or the human microbiome (Bacteroides, Streptococcus, Staphylococcus). In single cases, bacteria, Mycoplasma, fungi, and human cells were detected in the debris at the bottom of the storage tanks. In conclusion, the limited microbial load of the ice phase and in the debris of storage tanks can be effectively avoided by minimizing ice formation and by employing hermetically sealed sample containers.

The role of 'atypical' Brucella in amphibians: are we facing novel emerging pathogens?

AIMS: To discuss together the novel cases of Brucella infections in frogs with the results of published reports to extend our current knowledge on 'atypical' brucellae isolated from amphibians and to discuss the challenges we face on this extraordinary emerging group of pathogens. METHODS AND RESULTS: Since our first description, an additional 14 isolates from four different frog species were collected. Novel isolates and a subset of Brucella isolates previously cultured from African bullfrogs were characterized by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), Fourier transform-infrared (FT-IR) spectroscopy and broth microdilution susceptibility testing. MALDI-TOF MS worked very efficiently for an accurate bacterial identification to the genus level. Within the cluster analysis, 'atypical' brucellae grouped distant from Brucella melitensis and were even more separated by FT-IR spectroscopy with respect to their geographical origin. Minimum inhibitory concentrations of 14 antimicrobial substances are provided as baseline data on antimicrobial susceptibility. CONCLUSIONS: The case history of Brucella infections in amphibians reveals a variety of pathologies ranging from localized manifestations to systemic infections. Some isolates seem to be capable of causing high mortality in zoological exhibitions putting higher demands on the management of endangered frog species. There is considerable risk in overlooking and misidentifying 'atypical' Brucella in routine diagnostics. SIGNIFICANCE AND IMPACT OF THE STUDY: Brucella have only recently been described in cold-blooded vertebrates. Their presence in frog species native to Africa, America and Australia indicates a more common occurrence in amphibians than previously thought. This study provides an extensive overview of amphibian brucellae by highlighting the main features of their clinical significance, diagnosis and zoonotic potential.

Antimicrobial Susceptibility Patterns of Enterococcus faecalis and Enterococcus faecium Isolated from Poultry Flocks in Germany.

Between 2010 and 2011, 145 Enterococcus isolates (Enterococcus faecalis, n = 127; Enterococcus faecium, n = 18) were collected during routine bacteriologic diagnostics from broilers, layers, and fattening turkeys in Germany showing various clinical signs. The susceptibility to 24 antimicrobial agents was investigated by broth microdilution test to determine minimum inhibitory concentrations (MICs). All E. faecalis isolates (n = 127) were susceptible to the beta-lactam antibiotics ampicillin, amoxicillin-clavulanic acid, and penicillin. Corresponding MIC with 50% inhibition (MIC50) and MIC with 90% inhibition (MIC90) values of these antimicrobial agents were at the lower end of the test range (≤ 4 µg/ml). In addition, no vancomycin-resistant enterococci (VRE) were found. High resistance rates were identified in both Enterococcus species for lincomycin (72%-99%) and tetracycline (67%-82%). Half or more than half of Enterococcus isolates were resistant to gentamicin (54%-72%) and the macrolide antibiotics erythromycin (44%-61%) and tylosin-tartate (44%-56%). Enterococcus faecalis isolated from fattening turkeys showed the highest prevalence of antimicrobial resistance compared to other poultry production systems. Eighty-nine out of 145 Enterococcus isolates were resistant to three or more antimicrobial classes. Again, turkeys stood out with 42 (8 1%) multiresistant isolates. The most-frequent resistance patterns of E. faecalis were gentamicin, lincomycin, and tetracycline in all poultry production systems.

Bats and bacterial pathogens: a review.

The occurrence of emerging infectious diseases and their relevance to human health has increased the interest in bats as potential reservoir hosts and vectors of zoonotic pathogens. But while previous and ongoing research activities predominantly focused on viral agents, the prevalence of pathogenic bacteria in bats and their impact on bat mortality have largely neglected. Enteric pathogens found in bats are often considered to originate from the bats' diet and foraging habitats, despite the fact that little is known about the actual ecological context or even transmission cycles involving bats, humans and other animals like pets and livestock. For some bacterial pathogens common in human and animal diseases (e.g. Pasteurella, Salmonella, Escherichia and Yersinia spp.), the pathogenic potential has been confirmed for bats. Other bacterial pathogens (e.g. Bartonella, Borrelia and Leptospira spp.) provide evidence for novel species that seem to be specific for bat hosts but might also be of disease importance in humans and other animals. The purpose of this review is to summarize the current knowledge of bacterial pathogens identified in bats and to consider factors that might influence the exposure and susceptibility of bats to bacterial infection but could also affect bacterial transmission rates between bats, humans and other animals.