GUT MICROBIOTA CHARACTERIZATION AS A MARKER TO EVALUATE THE EFFECT OF DIFFERENT DIETARY REGIMES ON WATER BUFFALO CALVES DURING THE PREWEANING PERIOD

Growth and development of calves are highly influenced by the composition and activity of their associated gut microbiota. A classic example of the importance of the microbiota in ruminants is the rumen, where fermentation of dietary substrates due to bacteria results in the formation of short-chain fatty acids. Short-chain fatty acids are a major energy source for the host, and an important substrate for the development of the rumen epithelium. However, in newborn calves, milk is primarily digested in the small intestine, and microbes colonizing the small intestine can contribute to intestinal homeostasis, stimulation of the immune system, and enhancement of intestinal epithelium wellness and development. In particular the first three months of life result to be the most sensitive rearing period for the young calf. Calves are challenged by a series of stress factors after birth, including changes in their farming environment. Indeed, after birth, the living environment changes from the sterile uterus to natural outside conditions and, in addition, changes occur in nutrition, digestion and absorption from natural milk provided by the mother to feed that calves gain by themselves. Optimum level of nutrition in early life favors faster growth, earlier onset of puberty, enhanced productivity and colonization of gut microbiota, which can influence healthy status. Rearing healthy calves is very important as it can have a significant impact on their growth and milk production performance in adult life. Adequate calf development is therefore crucially important for the entire dairy industry. Pre-weaning calves makes them particularly vulnerable to specific diseases, such as enteritis, which, in this age group, is among the diseases causing the highest mortality rates. Colostrum quality, diet with formula milk and management, as well as calf-related hygiene practices (e.g., cleaning routine for feeding equipment, calving pens, group calf pens), can have a pronounced effect on calf health and mortality. One of the most important aspects is the establishing of a proper nutrition plan, which is fundamental for the development and health of calves. A correct and gradual transition from the neonatal phase to the subsequent phase of development ensures that the weaning phase is successfully overcome. In fact, during early life the diet may impact colonization of gut microbiota, which can influence health, leading to potential long-lasting consequences later in life. Today, there are still too many Italian farms exhibiting mortality rates higher than the physiological averages or that cannot sufficiently anticipate the weaning of calves to bring them at the next phase of growth, in a good state of health and with a rumen ready to digest fodder and concentrates,; this has led to a clear slowdown of genetic improvement in most farms. A functional diet chosen to promote the proper development of the gut microbiota is therefore a strategy to ensure animal welfare and productivity. In view of the commercial value of water buffalo milk, in addition, it has become necessary to optimize and simplify the weaning of water buffaloes calves, identifying protocols to replace breast milk without compromising the development and health of the animals themselves. This has led to use different types of feeding that consequently impacted on the development of the gastrointestinal microbiota and consequently on the health of animals, especially in calves, because in the first period of life the microbial colonization is not yet stable and therefore can be easily affected. Many studies have been conducted both in humans and in different animal species about the role of microbiota and how it changes in relation to diet. In particular, in the ruminants species, the attention was focused on the rumen microbiota, since rumen plays the main role in the digestion of the adult animal. In calves, instead, when the animal is monogastric, intestine is the site playing the most important role in the digestion so, it is important to focus attention on this organ and its microbial flora organization. Information is extremely limited on ruminant gut colonization, especially when focusing on the role of the microbiota in the early development of the gastro-intestinal (GIT) during the pre-ruminant period. Water buffalo farming is the main economic source for many families in many areas of the world: Europe, Australia, North America, South America and some African countries. For this reason it is important to invest in research in this area with the aim to integrated knowledge and to promote quality and innovation in primary productions. For these reasons the purpose of this study was to: - characterize the fecal microbiota in calves during the pre-weaning phase - determine which diet (water buffalo milk, formula milk or mixed diet) can contribute to the proper development of the gut microbiota - identify the most useful diet to promote the growth and development of calves. The results demonstrated the homogeneity in gut microbiota composition of newborn calves belonging to the same farm. In particular, consistent with what is described in dairy calves, the predominant phyla in newborn calves are: Proteobacteria, Firmicutes and Bacteroidetes. Differences are evident between samples collected at two time points and displaying maturation of the intestinal microflora in the first weeks of life. Difference occurs in groups feed with mixed diet were the microbiota structure is different and change the abundance of six genera in feces samples: Faecalibacterium, Clostridia_UCG-014, Bifidobacterium, Collinsella, Parabacteroides, Eubacterium_coprostanoligenes_group. The high-throughput sequencing demonstrated to be a suitable approach to the study of the gut microbiota of newborn calves allowing the characterization of the intestinal microbiota during the pre-weaning phase. The characterization of the gut microbiota in calves during the pre-weaning phase and monitoring changes over time could represent a useful tool for monitoring the health status of calves

Synthesis and Ring-Opening Metathesis Polymerization of a New Norbornene Dicarboximide with a Pendant Carbazole Moiety

A new norbornene dicarboximide presenting a pendant carbazole moiety linked by a p-methylene benzyl spacer is synthesized. This carbazole-functionalized monomer is polymerized via ring-opening metathesis polymerization using Grubbs third-generation catalyst. Microstructural analysis of resulting polymers performed by Nuclear Magnetic Resonance (NMR) shows that they are stereoirregular. Wide-angle X-ray diffraction (WAXD) and thermal (DSC) analysis indicate that polymers are also amorphous. With respect to the fluorescence analysis, both solution and film polymer samples exhibit only "normal structured" carbazole fluorescence, while excimer formation by overlap of carbazole groups is not detected

Complete Genome Sequencing of 10 Brucella abortus Biovar 3 Strains Isolated from Water Buffalo

Brucellosis is a zoonotic disease that affects both humans and animals. Its distribution is global, concentrated in the Mediterranean area, India, Central Asia, and Latin America. Here, we present a complete genome assembly of 10 Brucella abortus strains isolated from water buffaloes farmed in the Campania region of Italy

Complete Genome Sequencing of Eight Brucella abortus Biovar 1 Strains Isolated from Water Buffalo

Brucellosis is a zoonotic disease caused by bacteria of the genus Brucella The disease is endemic in many areas, causing chronic infections responsible for reproductive disorders in infected animals. Here, we present eight complete genome assemblies of eight Brucella abortus strains isolated from water buffaloes farmed in the Campania region

Whole-Genome Sequencing-Based Characterization of a Listeria monocytogenes Strain from an Aborted Water Buffalo in Southern Italy

Listeria monocytogenes is a Gram-positive pathogen causing life-threatening infections both in humans and animals. In livestock farms, it can persist for a long time and primarily causes uterine infections and encephalitis in farmed animals. Whole genome sequencing (WGS) is currently becoming the best method for molecular typing of this pathogen due to its high discriminatory power and efficiency of characterization. This study describes the WGS-based characterization of an L. monocytogenes strain from an aborted water buffalo fetus in southern Italy. The strain under study was classified as molecular serogroup IVb, phylogenetic lineage I, MLST sequence type 6, Clonal Complex 6, and cgMLST type CT3331, sublineage 6. Molecular analysis indicated the presence of 61 virulence genes and 4 antibiotic resistance genes. Phylogenetic analysis, including all the publicly available European L. monocytogenes serogroup IVb isolates, indicated that our strain clusterized with all the other CC6 strains and that different CCs were variably distributed within countries and isolation sources. This study contributes to the current understanding of the genetic diversity of L. monocytogenes from animal sources and highlights how the WGS strategy can provide insights into the pathogenic potential of this microorganism, acting as an important tool for epidemiological studies

Different Impacts of MucR Binding to the babR and virB Promoters on Gene Expression in Brucella abortus 2308

The protein MucR from Brucella abortus has been described as a transcriptional regulator of many virulence genes. It is a member of the Ros/MucR family comprising proteins that control the expression of genes important for the successful interaction of alpha-proteobacteria with their eukaryotic hosts. Despite clear evidence of the role of MucR in repressing virulence genes, no study has been carried out so far demonstrating the direct interaction of this protein with the promoter of its target gene babR encoding a LuxR-like regulator repressing virB genes. In this study, we show for the first time the ability of MucR to bind the promoter of babR in electrophoretic mobility shift assays demonstrating a direct role of MucR in repressing this gene. Furthermore, we demonstrate that MucR can bind the virB gene promoter. Analyses by RT-qPCR showed no significant differences in the expression level of virB genes in Brucella abortus CC092 lacking MucR compared to the wild-type Brucella abortus strain, indicating that MucR binding to the virB promoter has little impact on virB gene expression in B. abortus 2308. The MucR modality to bind the two promoters analyzed supports our previous hypothesis that this is a histone-like protein never found before in Brucella

A correlation of Mycobacterium bovis SB0134 infection between cattle and a wild boar (Sus Scrofa) in Campania region

A case of Mycobacterium bovis infection is described in a death adult female wild boar in the province of Avellino, Campania Region (Southern Italy). The carcass was sent to the Istituto Zooprofilattico Sperimentale del Mezzogiorno (IZSM) of Portici, Naples, Italy, where postmortem examination was performed. At necropsy, a disseminated granulomatous infection was observed, with involvement of various lymph node districts, spleen and lungs. Therefore, all lymph nodes were collected, together with spleen and lung lesions, in order to carry out bacteriological and molecular analyses that confirmed an uncommon disseminated Mycobacterium bovis infection. Subsequently, an analysis of the spoligotype, performed by the National Reference Center of Mycobacterium bovis in Brescia (Northern Italy), resulted in the spoligotype SB0134, previously identified in bovine outbreaks in the same area where the wild boar was found

Octupolar Metastructures for a Highly Sensitive, Rapid, and Reproducible Phage-Based Detection of Bacterial Pathogens by Surface-Enhanced Raman Scattering

The development of fast and ultrasensitive methods to detect bacterial pathogens at low concentrations is of high relevance for human and animal health care and diagnostics. In this context, surface-enhanced Raman scattering (SERS) offers the promise of a simplified, rapid, and high-sensitive detection of biomolecular interactions with several advantages over previous assay methodologies. In this work, we have conceived reproducible SERS nanosensors based on tailored multilayer octupolar nanostructures which can combine high enhancement factor and remarkable molecular selectivity. We show that coating novel multilayer octupolar metastructures with proper self-assembled monolayer (SAM) and immobilized phages can provide label-free analysis of pathogenic bacteria via SERS leading to a giant increase in SERS enhancement. The strong relative intensity changes of about 2100% at the maximum scattered SERS wavelength, induced by the Brucella bacterium captured, demonstrate the performance advantages of the bacteriophage sensing scheme. We performed measurements at the single-cell level thus allowing fast identification in less than an hour without any demanding sample preparation process. Our results based on designing well-controlled octupolar coupling platforms open up new opportunities toward the use of bacteriophages as recognition elements for the creation of SERS-based multifunctional biochips for rapid culture and label-free detection of bacteria