Genomic and phenotypic characterization of the Oropouche virus strain implicated in the 2022-24 large-scale outbreak in Brazil.

The Orthobunyavirus oropoucheense species encompasses a group of arthropod-borne zoonotic viruses transmitted by biting midges to animals including humans. Several large-scale human outbreaks caused by the prototype member of this species, Oropouche virus (OROV) have been documented since the 1970s and were primarily confined to the Amazon basin. However, since 2022, more widespread OROV outbreaks have been unfolding in Brazil and across South America, with cases exported to Cuba, Italy, Spain, USA and Germany. In Brazil, the virus has reached and established communitary transmission in all geographic areas of the country. We isolated, characterized the cytopathic effect and recovered the full genome of two OROV isolates from the 2022-24 outbreak detected in patients from the Pernambuco state. Phylogenetic data supports a direct introduction from the Amazonas state, the epicenter of the epidemics in the country. As case counts accumulate in the state mounting evidence is supporting the establishiment of sustained transmission chains. Continued studies are critical to understand the transmission cycle in this region, including the most important vectors and reservoirs, to appropriately deploy control measures.

Effects of supplementing direct-fed microbials on health and growth of preweaning Gyr × Holstein dairy calves.

This study aimed to evaluate the effects of direct-fed microbials (DFM) on health and growth responses of preweaning Bos indicus × Bos taurus (Gyr × Holstein) crossbred calves. Ninety newborn heifer calves (initial BW of 35 ± 4.0 kg) were used. At birth, calves were ranked by initial BW and parity of the dam and assigned to: (1) whole milk without DFM supplementation (CON; n = 30), (2) whole milk with the addition of 1.0 g/calf per day of a Bacillus-based DFM (BAC; n = 30), or (3) whole milk with the addition of 1.0 g/calf per day of BAC and 1.2 g/calf per day of Enterococcus faecium 669 (MIX; n = 30). Milk was fed individually during the study (77 d), and the BAC and MIX treatments were offered daily throughout the 77-d preweaning period. All calves were offered a starter supplement and corn silage starting on d 1 and 60 of age, respectively. Milk and starter supplement intake were evaluated daily, and BW was recorded on d 0 and at weaning (d 77). Diarrhea and pneumonia were assessed daily, and fecal samples were collected on d 0, 7, 14, 21, and at weaning (d 77) for assessment of the presence of bacterial and protozoal pathogens via qPCR. All data were analyzed using SAS (v. 9.4) with calf as the experimental unit and using single-df orthogonal contrasts (BAC + MIX vs. CON; BAC vs. MIX). Daily feeding of DFM, regardless of type, improved weaning BW. Odds ratio for occurrence of pneumonia was lower for DFM-supplemented calves, but its occurrence did not differ between BAC and MIX calves. No Salmonella spp. or Escherichia coli F41 were detected in any of the calves. The proportion of calves positive for E. coli F17 was greater for DFM calves on d 7 (92% and 96% vs. 81% for BAC, MIX, and CON, respectively), on d 21 (13% and 26% vs. 7% for BAC, MIX, and CON, respectively), and at weaning (48% and 35% vs. 22% for BAC, MIX, and CON, respectively). For Clostridium difficile, more DFM calves were positive on d 7 (65% and 30% vs. 35% for BAC, MIX, and CON, respectively) and 14 (20% and 28% vs. 7% for BAC, MIX, and CON, respectively), but proportion of positive calves was also greater for BAC versus MIX on d 7. More CON calves were positive for Clostridium perfringens on d 14 (14% vs. 3% and 8% for CON, BAC, and MIX, respectively) compared with DFM-fed calves. Incidence of calves positive for C. perfringens was greater in BAC than MIX on d 7 (50% vs. 18%), and greater for MIX than BAC at weaning (9% vs. 0%). For protozoa occurrence, a lower proportion of DFM calves were positive for Cryptosporidium spp. on d 7 (58% and 48% vs. 76% for BAC, MIX, and CON, respectively), but opposite results were observed on d 21 for Cryptosporidium spp. (3% and 11% vs. 0% for BAC, MIX, and CON, respectively) and Eimeria spp. on d 14 (7% and 8% vs. 0% for BAC, MIX, and CON, respectively) and 21 (50% and 59% vs. 38% for BAC, MIX, and CON, respectively). In summary, DFM feeding alleviated the occurrence of pneumonia and improved growth rates, while also modulating the prevalence of bacteria and protozoa in preweaning Gyr × Holstein calves.

Effects of supplementing a direct-fed microbial containing Enterococcus faecium 669 on performance, health, and metabolic responses of preweaning Holstein dairy calves.

This study aimed to evaluate the effects of Enterococcus faecium 669 supplementation on performance, health, parasitological, microbiological, and hematological responses of preweaning dairy calves. Forty-two newborn Holstein female calves (initial body weight [BW] 44 ± 4.5 kg) were used in the present study. At birth, calves were ranked by initial BW and assigned to 1 of 2 treatment groups: (1) whole milk (CON; n = 21) and (2) whole milk with the addition of direct-fed microbial Ent. faecium 669 (DFM; n = 21). During the entire experimental period (63 d), DFM was daily-fed at a rate of 2.5 × 109 cfu/head. All calves were offered a mixture of a starter feed and wheat straw for ad libitum consumption. Supplement intake was evaluated daily, whereas calves were weighed on a weekly basis from d 0 to weaning (d 63). Diarrhea was assessed once a day, and fecal and blood samples were collected for microbiological, parasitological, and hematological responses. All data were analyzed with SAS using calf as the experimental unit. A treatment × week interaction was observed for BW, as DFM-supplemented calves were heavier than CON cohorts on d 56 (+ 4.7 kg) and at weaning on d 63 (+ 4.8 kg). A similar interaction was observed for average daily gain (ADG) and dry matter intake (DMI), with greater ADG for DFM-supplemented calves from d 35 to 42, greater ADG and DMI from d 49 to 56, and greater DMI from d 56 to weaning. Moreover, diarrhea occurrence tended to be lower, whereas rectal temperature was 0.2°C lower for DFM-supplemented calves. Treatment × day interactions were observed for the occurrence and counts of Eimeria spp., as DFM-supplemented calves tended to have a reduced number of positive observations on d 42 of the study versus CON, and a significant reduction in positive animals from d 21 to 42 was observed in the DFM group but not in CON calves. For Cryptosporidium spp., no treatment effects were observed on overall occurrence (%), but DFM-supplemented calves had a greater count of oocyst per gram versus CON. No treatment × day interaction or main treatment effects were observed for any of the blood variables analyzed herein, exception being monocytes concentration. In summary, preweaning Ent. faecium 669 supplementation improved performance, diarrhea occurrence, and reduced the number of calves positively-detected for Eimeria spp.

Utilizing intramammary Melaleuca alternifolia as an organic internal sealant for dry-off therapy in Murrah buffaloes.

The effects of intramammary dry cow therapy based on the administration of 5% Melaleuca alternifolia tea tree essential oil (TTO) as an internal teat sealant to Murrah cows were evaluated. A longitudinal prospective and retrospective negative control study was performed using 12 buffaloes from a total of 20 Murrah buffaloes on an organic farm, with the cow used as a control for herself. The minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) for treatments with pure oil (TTO) and medication containing 5% TTO (O5) were determined. The buffaloes were clinically examined, and the teats were evaluated using thermography and ultrasound. Udder health was monitored during the first 100 days in milk (DIM) using milk somatic cell count (SCC) and California mastitis test (CMT). Laboratory tests against standard strains Staphylococcus aureus ATCC®25,923™, Escherichia coli ATCC®25,922™, and wild bacterial strains showed maximum MIC values of 50 µL/mL for the TTO and O5 treatments. One wild-type S. aureus strain showed no MBC. No adverse effects were observed after the intramammary application of TTO. The CMT and SCC values were similar (P > 0.05) for all observations. The medication containing 5% TTO was effective in vitro and compatible with the intramammary tissue in vivo of Murrah buffaloes. TTO was safe, not inducing inflammatory processes or other modifications of the teat detectable by thermography or ultrasound. It was able to protect buffaloes during the dry period under field conditions, demonstrating potential use as a teat sealant for organic farms.

Comparison between MIBI-based radiopharmaceuticals for parathyroid scintigraphy: quantitative evaluation and correlation with clinical-laboratory parameters.

OBJECTIVE: Parathyroid scintigraphy is mandatory for the identification of hyperfunctioning parathyroid glands in hyperparathyroidism (HPT). The use of 99mTc-methoxy-isobutyl-isonitrile (MIBI) as radiopharmaceutical for parathyroid scintigraphy is considered the most valid and useful considering its uptake mechanism. Several MIBI-based radiopharmaceuticals are commercially available (i.e., MediMIBI, TechneMIBI, Stamicis). They seem to have similar physico-chemical characteristics and the choice between them is based on commercial criteria, even though some differences in qualitative scintigraphic results have been appreciated. Aims of the study were: first, to compare the scintigraphic quantitative data of MediMIBI, TechneMIBI, and Stamicis, particularly in the view of a personalized medicine approach; second, to investigate the potential effect of clinical-laboratory data on image quality using one of these radiopharmaceuticals. METHODS: Patients with diagnosis of HPT, who underwent a parathyroid scintigraphy using one of the three MIBI-based radiopharmaceuticals between December 2018 and October 2020, have been retrospectively identified. Parameters derived from regions of interest (ROIs) drawn on three different sites were obtained: a reasonable parathyroid lesion detected, an area in the lateral neck considered as the background, and the hepatic dome as the site of MIBI physiological uptake. Laboratory and clinical data, such as serum calcium, PTH, vitamin D, and creatinine levels, as well as possible drug-mediated interferences were considered. RESULTS: Among 250 patients included, 83 (33.2%) had the parathyroid scintigraphy using MediMIBI, 84 (33.6%) using TechneMIBI and 83 (33.2%) using Stamicis. The ROIs on the parathyroid uptake at early images, on the background, and on the hepatic dome were statistically different among the three groups (p < 0.05). No significant differences were found in the remaining semi-quantitative parameters among the three groups, not even considering clinical-laboratory data. CONCLUSIONS: Some differences in semi-quantitative parameters emerged among MIBI-based radiopharmaceuticals for parathyroid scintigraphy. This might justify the different qualitative scintigraphic results obtained using one or another of the cited radiopharmaceuticals.

ConfID: an analytical method for conformational characterization of small molecules using molecular dynamics trajectories.

MOTIVATION: The conformational space of small molecules can be vast and difficult to assess. Molecular dynamics (MD) simulations of free ligands in solution have been applied to predict conformational populations, but their characterization is often based on clustering algorithms or manual efforts. RESULTS: Here, we introduce ConfID, an analytical tool for conformational characterization of small molecules using MD trajectories. The evolution of conformational sampling and population frequencies throughout trajectories is calculated to check for sampling convergence while allowing to map relevant conformational transitions. The tool is designed to track conformational transition events and calculate time-dependent properties for each conformational population detected. AVAILABILITY AND IMPLEMENTATION: Toolkit and documentation are freely available at http://sbcb.inf.ufrgs.br/confid. CONTACT: marcelo.poleto@ufv.br or bigrisci@inf.ufrgs.br. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Anticancer activity of monoterpenes: a systematic review.

Secondary metabolites have been recognized for centuries as medicinal agents, in particular monoterpenes which have been the target of research in the discovery of antineoplastic drugs, as they have potential antitumor effect and low toxicity and are used as additives in foods and cosmetics. Another advantage of monoterpenes is structural diversity, which gives greater plasticity when interacting with cells. The purpose of this review was to summarize and critically discuss the anticancer potential of monoterpenes and their respective mechanisms of action. A systematic review of articles in the MEDLINE/PubMed, Web of Science, Scopus and Science Direct electronic databases was independently conducted by three reviewers using the combination of the following keywords: monoterpenes AND anticancer AND in vitro. Restriction in selecting articles followed pre-established inclusion and exclusion criteria by the reviewers, and also a time limitation with works published between 2015 and 2019 being selected. In total, 39 works were deemed eligible for inclusion in the final review. Monoterpenes have cytotoxic activity in a wide variety of tumor cell lines, and mainly appear to exert this effect by inducing apoptosis caused by oxidative stress. In addition, improved use of monoterpenes when used in drug delivery systems and the synergistic effect with conventional chemotherapeutic drugs are reported. These findings validate this class of compounds as a promising source of chemotherapeutic drugs yet to be explored.

Parathyroid scintigraphy in primary hyperparathyroidism: comparison between double-phase and subtraction techniques and possible affecting factors.

PURPOSE: Parathyroid scintigraphy is superior to other imaging techniques in detecting hyperfunctioning parathyroid glands. It is mainly performed using double-phase or dual-tracer subtraction methods. Neither of the techniques is perfect and different protocols are being used. We aimed to evaluate the accuracy of double-phase and subtraction methods in detecting abnormal gland as well as the potential effects of coexisting thyroid disease and clinical-laboratory data. METHODS: We considered patients with primary hyperparathyroidism who underwent parathyroid surgery, after a parathyroid scintigraphy between April 2015 and February 2017. Sixty-eight patients were included; in 45 cases (66.2%), a thyroid disease was coexistent. Diagnostic performances of the two techniques were compared. The effect of thyroid disease and clinical-pathological data on examination interpretation was considered. RESULTS: Double-phase scintigraphy showed higher sensitivity and accuracy in detecting the exact abnormal gland compared to the digital subtraction (90% and 75% vs. 76% and 66%, respectively). For double-phase technique, sensitivity and accuracy were higher in cases with no thyroid disease when compared to those with thyroid disease (92% and 86% vs. 88% and 69%, respectively). Similarly, for digital subtraction, sensitivity and accuracy were higher in the absence of thyroid disease compared to their presence (84% and 79% vs. 70% and 58%, respectively). There was no significant variation in the performance of both techniques, considering clinical-laboratory data. CONCLUSIONS: Double-phase scintigraphy has been more accurate than digital subtraction. The presence of thyroid disease could be a possible limit, affecting the subtraction more than the double-phase technique. Clinical data did not influence the scintigraphic outcome.

Tuning the biomimetic performances of 4-hydroxyproline-containing cyclic peptoids.

Five new cyclic peptoids containing (2S,4R)-4-hydroxyproline (Hyp) residues have been designed and synthesized using a mixed "submonomer/monomer" approach. Alkali metal cation affinities and ion transport activities were assessed by experimental (NMR and HPTS assay in liposomes) and computational methods. Easy functionalization of hydroxyproline residues afforded a bouquet of cyclic oligomers showing correlation between ion transport abilities and cytotoxic activities on selected human cancer cell lines.

Short communication: a novel multispecies bacteria-based direct-fed microbial supports in vitro gut barrier integrity challenged with a pathogen or pro-inflammatory cytokines.

We conducted two experiments to evaluate the effects of a novel bacterial-based direct-fed microbial (DFM) on intestinal barrier integrity using the in vitro transepithelial electrical resistance (TEER) assay. In experiment 1, human-derived Caco-2 cells received or not (CON) a DFM containing Ligilactobacillus (formerly Lactobacillus) animalis 506, Propionibacterium freudenreichii 507, Bacillus paralicheniformis 809, and B. subtilis 597 (BDP; BOVAMINE DEFEND® Plus) at a rate of 1 × 108 CFU/transwell. Concurrently with treatment application (CON or BDP), a pathogenic challenge of Clostridium perfringens type A was added alone (PAT) or with BDP (PAT + BDP) at a rate of 2.8 × 107 CFU/transwell in a 2 × 2 factorial arrangement. In experiment 2, Caco-2 cells were also assigned in a 2 × 2 factorial design to CON or BDP and then, 2 h post-treatment administration (CON and BDP), a mixture of tumor necrosis factor-alpha (TNF-α) and interferon-gamma (IFN-γ) was added alone (CYT) or with BDP (CYT + BDP) at a 10:1 ratio, respectively. In both experiments, TEER was measured for 18 h. In experiment 1, a DFM × pathogen × hour interaction was observed for TEER (P < 0.0001). Adding the PAT alone initially tended to increase TEER vs. CON from 1.1 to 2.2 h (P ≤ 0.09), increased TEER at 3.2 h (P < 0.01), but reduced TEER from 5.4 to the end of the experimental period at 18.4 h (P ≤ 0.01). On the other hand, adding DFM, with or without the pathogenic challenge, yielded greater TEER vs. CON-CON and CON-PAT for most of the experimental period (P ≤ 0.04). A similar interaction was detected and reported in experiment 2 (P < 0.0001). The CYT challenge reduced mean TEER compared with all other treatments from 3.2 h to the remainder of the study (P ≤ 0.03). On the other hand, BDP-CYT was able to maintain the integrity of the epithelial cells when compared with CON-CON throughout the experimental period (P ≤ 0.03), the exception being at 3.2 h (P = 0.20). Moreover, BDP-CON increased (P ≤ 0.04) TEER when compared with CON-CON from 3.2 to 18.4 h, but also in comparison with BDP-CYT from 4.3 to 18.4 h post-DFM and challenge administration into the cells. In summary, C. perfringens type A and a pro-inflammatory cytokine cocktail compromised the integrity of intestinal epithelial cell monolayers in vitro, whereas adding a multispecies bacteria-based DFM counteracted these damaging effects.

Two experiments were designed to evaluate the effects of adding a bacterial-based direct-fed microbial (DFM) containing Lactobacillus animalis 506, Propionibacterium freudenreichii 507, Bacillus paralicheniformis 809, and Bacillus subtilis 597 on the integrity of intestinal epithelial cells challenged with Clostridium perfringens type A or a pro-inflammatory cytokine cocktail. Regardless of the challenge, the addition of the DFM maintained the integrity of the intestinal epithelial cells in vitro. These results help to elucidate the potential beneficial effects that the bacterial-based DFM containing L. animalis 506, P. freudenreichii 507, B. paralicheniformis 809, and B. subtilis 597 may bring to livestock species.

Bacillus paralicheniformis 809 and Bacillus subtilis 810 support in vitro intestinal integrity under hydrogen peroxide and deoxynivalenol challenges.

We designed and conducted two in vitro experiments to evaluate the effects of two Bacillus spp. probiotics on gut barrier integrity using the transepithelial electrical resistance (TEER) assay under two different challenge models. In Exp. 1, intestinal epithelial cells received or not (CON) B. paralicheniformis 809 (BLI) or B. subtilis 810 (BSU) at a rate of 1 × 108 colony forming units (CFU)/transwell. Two hours after treatment application (CON, BLI, or BSU), 5 mM of the reactive oxygen species hydrogen peroxide, mimicking mucosal oxidative stress, was added alone (HYP) or with each of the Bacillus spp. (HYP + BLI or HYP + BSU). In Exp. 2, cells were assigned to the same treatments as in Exp. 1 (CON, BLI, and BSU), or mycotoxin deoxynivalenol (DON), which was added alone or in combination with BLI or BSU, resulting in another two treatments (DON + BLI and DON + BSU). Transepithelial electrical resistance was measured for 14 h postchallenge. In Exp. 1, a treatment × hour interaction was observed for TEER (P < 0.0001). Adding BLI and BSU resulted in greater TEER values vs. CON for most of the experimental period (P < 0.02), whereas HYP reduced mean TEER and area under the curve (AUC), while increasing the amount of sugar that translocated through the monolayer cells (P < 0.001). A treatment × hour interaction was also observed in Exp. 2 (P < 0.0001), as DON led to an immediate and acute drop in TEER that lasted until the end of the experimental period (P < 0.0001). Both BLI and BSU alleviated the DON-induced damaging effects on the integrity of intestinal epithelial cells, whereas both Bacillus spp. alleviated the damage caused by DON alone and the proportion of sugar that translocated through the monolayer cells was not different between CON and DON + BLI (P = 0.14) and DON + BLI and DON + BSU (P = 0.62). In summary, both Bacillus spp. strains (B. paralicheniformis 809 and B. subtilis 810) were able to counteract the damaging effects of the challenge agents, hydrogen peroxide and deoxynivalenol, on gut barrier integrity.

Effects of a Bacillus-based direct-fed microbial on performance, blood parameters, fecal characteristics, rumen morphometrics, and intestinal gene expression in finishing beef bulls.

We evaluated the effects of supplementing direct-fed microbials (DFM), containing Bacillus licheniformis and Bacillus subtilis, on performance, rumen morphometrics, intestinal gene expression, and blood and fecal parameters in finishing bulls. Nellore × Angus bulls (n = 144; initial BW = 401 ±â€…45.5 kg) were distributed at random in 36 pens (4 bulls/pen and 18 pens/treatment), following a completely randomized design. A ground corn-based finishing diet was offered for ad libitum intake twice a day for 84 d, containing the following treatments: 1) control (without DFM); 2) DFM (B. licheniformis and B. subtilis) at 6.4 × 109 CFU (2 g) per animal. The data were analyzed using the MIXED procedure of SAS, with a pen representing an experimental unit, the fixed effect of the treatment, and the random effect of pen nested within the treatment. For fecal parameters (two collections made), the collection effect and its interaction with the treatment were included in the model. Bulls that received the DFM had a decreased dry matter intake (P ≤ 0.01), did not differ in average daily gain (2.05 kg; P = 0.39), and had a 6% improvement in gain:feed (P = 0.05). The other performance variables, final BW, hot carcass weight, and hot carcass yield, did not differ (P > 0.10). Plasma urea-N concentration decreased by 6.2% (P = 0.02) in the bulls that received DFM. Glucose, haptoglobin, and lipopolysaccharides were not different between treatments (P > 0.10). Ruminal morphometrics were not affected by the treatment (P > 0.10). The use of DFM tended to reduce fecal starch (P = 0.10). At slaughter, bulls fed DFM had an increased duodenal gene expression of tryptophan hydroxylase-1 (P = 0.02) and of superoxide dismutase-1 (P = 0.03). Overall, supplementation with DFM based on B. licheniformis and B. subtilis to Nellore × Angus bulls in the finishing phase decreased dry matter intake, did not influence ADG, improved gain:feed, and increased the expression of genes important for duodenal function.

One of the main alternatives of additives to modulate the microbial population in the gastrointestinal tract (GIT), especially in the intestine, is the use of direct-fed microbials (DFM). This class of additives comprises all the feed products that contain a live or naturally occurring source of microorganism. The inclusion of DFM in diets of ruminants in the finishing phase may improve gain:feed by modifying the composition of the microbial community in the GIT to bring about a better symbiotic relationship with the host. These effects may be achieved with the use of Bacillus spp. bacteria, such as Bacillus licheniformis and Bacillus subtilis. Mixtures of these bacteria are able to foster positive effects in the finishing phase of beef cattle fed high-energy diets, which reinforces the need for studies that examine the effects and mechanisms of these species. In this study, feedlot Nellore × Angus bulls that received a DFM composed of B. licheniformis and B. subtilis had decreased dry matter intake, no influence on average daily gain, improved gain:feed, and an increase in expression of genes important for duodenal function.

Supplementing a Bacillus-based direct-fed microbial improves feed efficiency in lactating dairy cows.

This experiment was conducted to evaluate the effects of feeding a Bacillus-based direct-fed microbial (DFM) on performance and nutrient digestibility of lactating dairy cows. Seventy-six lactating (42 ±â€…6 days in milk [DIM]) Holstein-Friesian primiparous and multiparous cows were enrolled to a 16-wk experiment. Cows were blocked by lactation number and DIM and within blocks, assigned to 1 of the 2 treatments: 1) basal partial-mixed ration (PMR) without DFM addition (n = 38; CON) or 2) basal PMR with the addition of 3 g/head/d of a DFM containing B. licheniformis 809 and B. subtilis 810 (n = 38; BOVACILLUS, Chr. Hansen A/S, Hørsholm, Denmark; DFM). The DFM was mixed in a protein-based pellet, whereas the CON group was fed the same pellet without DFM (0.6 kg/cow/d). The PMR contained (dry matter [DM] basis) 50% of forage and 48% of a concentrate feed based on corn meal, soybean meal, wheat meal, wheat middlings, and a mineral-vitamin premix, with the remaining part of the diet being represented by the pellet used as a carrier for the treatments (CON and DFM). Dry matter intake (DMI), milk yield, and production efficiency were recorded daily, whereas milk protein and fat concentrations were recorded using electronic milk meters. An additional milk sample was collected every second week of the study for milk composition. On week 15 of the study, fecal samples were collected from each cow for apparent nutrient digestibility calculation. All data were analyzed using the MIXED procedure of SAS (version 9.4; SAS Inst. Inc., Cary, NC). No treatment effects were observed on cow final body weight, daily DMI, milk yield, energy-corrected milk (ECM), ECM efficiency, milk composition (yield or content), and somatic cell count (SCC) (P ≥ 0.12). However, cows fed DFM had a greater feed and N efficiency (P ≤ 0.03) compared to cows fed CON. Moreover, DM digestibility tended to be greater for DFM-fed cows when compared to CON (P = 0.10), whereas no further nutrient digestibility differences were observed (P ≥ 0.24). In summary, supplementing a DFM containing Bacillus licheniformis and B. subtilis benefited feed efficiency of lactating dairy cows fed a PMR, while also tending to improve the digestibility of DM.

Effects of supplementing a Bacillus-based direct-fed microbial on performance, nutrient digestibility, rumen fermentation characteristics, and metabolic responses of lactating dairy cows.

This experiment was conducted to evaluate the effects of feeding a Bacillus-based direct-fed microbial (DFM) on performance, nutrient digestibility, rumen fermentation, and metabolic response of lactating dairy cows. Sixty-eight lactating (50 ± 6 d in milk) Holstein-Friesian (n = 20) and Swedish Red (n = 48) cows were enrolled to a 15-wk experiment. Cows were blocked by breed, lactation number, and days in milk and, within blocks, assigned to 1 of the 2 treatments: (1) basal partial mixed ration (PMR) without DFM (n = 34; CON) or (2) basal PMR with the addition of 3 g/head per day of a DFM containing Bacillus licheniformis and Bacillus subtilis (n = 34; DFM). The DFM was mixed in a protein pellet, whereas the CON group was fed the same pellet without DFM (1 kg/cow per day). The PMR contained 53% clover grass silage and 47% compound feed plus 3 kg of a concentrate (dry matter basis) offered during milking. Milk yield and production efficiency were recorded daily, whereas milk samples were collected for 24 h every second week of the study for milk composition. During the experimental period, fecal, rumen fluid, and blood samples were collected from each cow for apparent nutrient digestibility, rumen fermentation, and metabolic responses, respectively. All data were analyzed using the MIXED procedure of SAS. No treatment effects were observed on cows final body weight and daily dry matter intake. However, cows fed DFM had a greater milk yield, milk production efficiency, lactose and total solids yield, and also tended to have a greater energy-corrected milk production efficiency and milk protein yield. No significant differences were observed on nutrient digestibility and total volatile fatty acids, but molar proportion of acetate was greater for cows fed DFM. In contrast, molar proportion of propionate was greater and butyrate tended to be greater for CON. Cows fed DFM had greater mean plasma insulin-like growth factor-I (IGF-I), but no differences were observed for plasma glucose and insulin. In summary, supplementing a Bacillus-based DFM benefited productive responses of lactating dairy cows, while also modulating rumen fermentation and serum IGF-I.

Short communication: A multispecies bacterial-based direct-fed microbial alleviates Salmonella invasion and supports in vitro epithelial integrity.

Managing bacterial infections is of great importance in livestock production, particularly those caused by Salmonella enterica serovars Typhimurium or Dublin, which can impact both animal health and performance, as well as human food safety. Direct-fed microbials (DFM) can support gastrointestinal function and alleviate the potential negative effects of bacterial infections. In the present study, the capacity of a multispecies bacterial-based DFM containing Ligilactobacillus (formerly Lactobacillus) animalis 506, Propionibacterium freudenreichii 507, Bacillus licheniformis 809, and B. subtilis 597 to reduce S. Typhimurium ATCC14028 invasion was investigated using a co-incubation model with the HT29-MTX-E12 cell line (Exp. 1). Next, a possible antagonistic effect of the DFM against S. Dublin ATCC 41286 was evaluated using an in vitro agar well diffusion method following a co-incubation of 48 h (Exp. 2). At last, a series of experiments were performed to evaluate how different doses (6.25 × 106, 2.50 × 107, or 1.00 × 108 CFU/well) of the DFM would support the integrity of intestinal epithelial cells challenged or not with S. Typhimurium ATCC14028 or hydrogen peroxide under a transepithelial electrical resistance (TEER) assay with Caco-2 cells (Exp. 3 and 4). In Exp. 1, BDP significantly (P < 0.001) reduced by 90.8% the invasion of S. Typhimurium into HT29-MTX-E12 cells, whereas viability of the potentially harmful bacteria was reduced by 21.0% (P < 0.0001). In Exp. 2, the antagonistic properties of BDP towards S. Dublin were confirmed by the detection of a clear inhibition zone (size = 8.6 mm). Lastly, without challenge, the lowest dose of the DFM (6.25 × 106 CFU) provided the greatest support on the cells (treatment × hour; P < 0.0001). However, when the cells were challenged with S. Typhimurium, all doses alleviated the loss of integrity caused by the pathogen (treatment × hour; P < 0.0001). In cells challenged with hydrogen peroxide, the greater dose (1.00 × 108 CFU) supported the cells for a longer period of time (treatment × hour; P < 0.0001). These in vitro findings set the stage for exploring the potential benefits of using a novel DFM as a promising tool and strategy to mitigate Salmonella enterica infections in ruminants and improve animal health, food safety, and public health. Further in vivo confirmation needs to be developed to validate these preliminary in vitro results.

Supplementing a Bacillus-based probiotic to high-risk stocker cattle.

This experiment evaluated the performance, health, and physiological responses of high-risk steers receiving a Bacillus-based probiotic during a 90-d grazing period. A total of 240 Angus-influenced steers were used in this experiment that was replicated over 2 yr (120 steers/year). Each year, steers were obtained from an auction yard and transported to the experimental facility (120 km). Steer body weight (BW) was recorded at arrival (day -1), and this value was averaged with BW recorded on day 0 to represent the initial BW (236.6 ± 1.5 kg). On day 0, steers were ranked by BW and allocated to 1 of 12 pastures with stockpiled native grass (4-ha pastures; 10 steers/pasture). Pastures were randomly assigned to receive daily supplementation with dried distillers' grains at 1% of BW containing either: 1) Bacillus subtilis + B. licheniformis probiotic (BOV; 2 g/steer daily of Bovacillus; Novonesis, Horsholm, Denmark) or 2) no feed additive (CON). Cattle received treatments from days 0 to 90, in addition to free-choice access to water and mineral + vitamin mix without ionophore. Steers were assessed for bovine respiratory disease (BRD) signs daily. Blood samples were collected and full BW was recorded on days 0, 14, 28, 56, and 90. Shrunk BW was recorded on day 91 after 16 h of feed and water restriction, and a 4% pencil shrink was used to calculate the final BW. Average daily gain (ADG) was calculated based on initial and final BW. No treatment effects were detected (P ≥ 0.73) for steer final BW and ADG. A treatment × day interaction was detected (P ≤ 0.05) for plasma haptoglobin concentration, which was greater for CON steers on days 14 and 28 (P ≤ 0.02). Incidence of BRD signs did not differ (P = 0.97) between treatments (51.7% and 51.3% for BOV and CON, respectively; SEM = 7.70). However, steer mortality + removals for health complications were greater (P = 0.01) in CON compared to BOV (0.00% vs. 5.04%, respectively; SEM = 1.41). Supplementing BOV improved (P ≤ 0.04) total pasture-based liveweight change (643 vs. 502 kg/pasture, respectively; SEM = 45) and final pasture-based total liveweight (3,007 vs. 2,869 kg/pasture, respectively; SEM = 46). Collectively, supplementation with a probiotic based on B. subtilis and B. licheniformis to high-risk stocker cattle did not alleviate the incidence of BRD signs nor improved ADG, but decreased acute-phase protein response, reduced steer mortality + removal, and increased pasture-based productivity during a 90-d grazing period.

Stocker cattle are exposed to several stressors within a short period of time, which impair their immunity and lead to bovine respiratory disease (BRD). With the increased regulations regarding the use of antimicrobials in cattle nutrition, novel dietary strategies to improve health and productivity of stocker cattle are warranted. One example is supplementing Bacillus-based probiotics, which promote performance and immunity in high-stress cattle. In this study, steers were purchased from a commercial auction yard soon after weaning, transported to the research facility, and assigned initial processing within a 48-h period. Steers were assigned to pastures and were supplemented or not with the Bacillus-based probiotic during a 90-d grazing period. In general, supplementing steers with the Bacillus-based probiotic did not impact growth rates or BRD incidence. However, no steers that received the Bacillus-based probiotic died from BRD consequences nor were removed from the experiment due to health reasons, whereas 5% of unsupplemented steers did not complete the 90-d experiment. Consequently, pasture-based liveweight gain was increased by 28% due to Bacillus-based probiotic supplementation. Results from this study indicate that supplementing a B. subtilis + B. licheniformis probiotic could be an alternative to improve the health and overall productivity of high-risk stocker cattle.

Effects of feeding an inoculated corn silage with or without a direct-fed microbial on dry matter intake, milk production, and nutrient digestibility of high-producing lactating Holstein cows.

This study evaluated the effects of inoculating corn silage and/or feeding a direct-fed microbial (PRO) on performance and nutrient digestibility of lactating dairy cows. At harvesting, corn silage was treated either with water (culated or not [CON]) or Lactococcus lactis and Lentilactobacillus buchneri (INC; SiloSolve FC) at 1.5 × 105 cfu/g of corn silage. Ten mini silos and one farm-scale silo bunker per treatment were prepared for the laboratory and the lactating dairy cow trial, respectively. Five mini silos per treatment were opened on days 2 or 90 post-ensiling for pH measurement, as well as chemical analysis and aerobic stability, respectively. The farm-scale silo bunkers were opened 77 d post-ensiling for the beginning of the lactating cow trial. Eighty lactating Holstein cows were assigned in a 2 × 2 factorial design to: (1) CON silage without PRO (CON-CON; n = 20), (2) CON silage with PRO at 14 g/head/d (CON-PRO; n = 20), (3) INC silage without PRO (INC-CON; n = 20), and (4) INC silage with PRO at 14 g/head/d (INC-PRO; n = 20). Concurrently with the feeding trial, eight cows per treatment were chosen for nutrient digestibility. The pH of the corn silage was not affected by the silage inoculant (P ≥ 0.29), but INC yielded greater concentration of acetic acid and longer aerobic stability (P < 0.01). Dairy cows fed INC had a lower mean total dry matter intake (DMI), milk protein content, and somatic cell counts vs. CON (P ≤ 0.02). On the other hand, milk and fat- and protein-corrected milk (FPCM) production efficiency, milk urea-N, DM, crude protein, and starch digestibility were greater for INC-fed cows (P ≤ 0.03). Feeding direct-fed microbials (DFM) improved mean body weight, milk yield, and FPCM, as well as milk protein and lactose yield (P ≤ 0.05), but reduced milk fat and protein content (P = 0.02). A silage inoculant × DFM interaction was observed for milk production efficiency, milk protein and lactose content, and somatic cell count (P ≤ 0.05). Dairy cows fed INC-CON had a greater milk production efficiency and milk lactose content (P ≤ 0.04), but INC-PRO had lower milk protein content and SCC (P ≤ 0.03). In summary, inoculating L. lactis and L. buchneri increased acetic acid content and aerobic stability of corn silage, reduced DMI, but improved milk production efficiency and nutrient digestibility of lactating Holstein dairy cows. On the other hand, feeding PRO improved milk, protein, and lactose yield. Additionally, combining the feeding of an inoculated corn silage with PRO reduced milk somatic cell count.

Maternal pre- and postpartum supplementation of a Bacillus-based DFM enhanced cow and calf performance.

This study evaluated the effects of maternal supplementation of a Bacillus-based direct-fed microbial (DFM) on the physiology and growth performance of Bos indicus-influenced cow-calf pairs. On day 0 (~139 d before expected calving date), 72 fall-calving, Brangus crossbred beef heifers (20 to 22 mo of age) pregnant with first offspring were stratified by their initial body weight (BW; 431 ±â€…31 kg) and body condition score (BCS; 6.0 ±â€…0.36; scale 1 to 9), and randomly allocated into 1 of 12 bahiagrass pastures (1 ha and six heifers per pasture). Treatments were randomly assigned to pastures (six pastures per treatment) and consisted of heifers supplemented with 1 kg/d of soybean hulls (dry matter, DM) that was added (BAC) or not (CON) with DFM containing Bacillus subtilis and B. licheniformis (Bovacillus; Chr. Hansen A/S, Hørsholm, Denmark). Treatments were provided from days 0 to 242 (139 ±â€…4 d prepartum to 104 ±â€…4 d postpartum). Calves were weaned on day 242 (96 ±â€…30 d of age) and then allocated into 1 of 16 drylot pens and fed the same concentrate at 3.25% of BW (DM) until day 319. Maternal treatment effects were not detected (P ≥ 0.29) for herbage allowance and forage chemical composition. Heifer BCS on days 39 and 63 tended (P ≤ 0.09) to be greater for BAC vs. CON heifers, whereas heifer BCS on day 91 was greater (P = 0.01) for BAC vs. CON heifers. Heifer BCS did not differ (P ≥ 0.20) between treatments on days 179 and 242. Plasma glucose concentration did not differ from days 0 to 63 (P ≥ 0.14) but were greater (P < 0.01) on day 179 and tended (P = 0.09) to be greater on day 242 for BAC vs. CON heifers. Calf BW at birth, ADG from birth to weaning, and BW at weaning did not differ (P ≥ 0.19) between treatments, but calf BW at drylot exit (day 319) was greater (P = 0.05) for BAC vs. CON calves. Maternal treatment effects were not detected (P ≥ 0.42) for calf serum concentration of IgG at birth and postvaccination plasma concentrations of glucose, cortisol, and haptoglobin. Serum titers against bovine respiratory syncytial virus (BRSV) were greater (P = 0.04) for BAC vs. CON calves on day 287, whereas seroconversion against parainfluenza-3 virus (PI-3) was greater (P < 0.01) for BAC vs. CON calves on day 271. Thus, maternal supplementation of a Bacillus-based DFM increased prepartum BCS gain and postpartum plasma glucose concentration of heifers and led to positive carryover effects on postweaning BW gain and humoral immune response in their offspring.

Direct-fed microbials (DFM), such as Bacillus spp., have been shown to produce a wide variety of enzymes related to nutrient digestion and to support gastrointestinal tract immune function and integrity, leading to increased nutrient digestibility and cattle performance. Nutritional management of beef cows during gestation and early lactation has been associated with enhanced future offspring growth performance and immune response following birth. The present study combined the use of Bacillus-based DFM for pregnant heifers during critical production stages (late gestation and early lactation) to promote the overall performance of heifers and their offspring. Heifers offered Bacillus-based DFM had greater body condition score at calving and postpartum plasma concentration of glucose, whereas their offspring had similar body weight at birth, but greater growth performance when fed relatively high amounts of protein and energy in drylot compared to cohorts born from heifers that did not receive Bacillus-based DFM supplementation.

Influence of BOVAMINE DEFEND Plus on growth performance, carcass characteristics, estimated dry matter digestibility, rumen fermentation characteristics, and immune function in finishing beef steers.

One hundred and eighty crossbred beef steers (406.0 ±â€…2.2 kg) were used to determine the impact of a novel direct-fed microbial (DFM) on growth performance, carcass characteristics, rumen fermentation characteristics, and immune response in finishing beef cattle. Steers were blocked by body weight (BW) and randomly assigned, within block, to 1 of 2 treatments (3 replicates/treatment: 30 steers/replicate). Treatments included: (1) no DFM (control) and (2) DFM supplementation at 50 mg ∙ animal-1 ∙ d-1 (BOVAMINE DEFEND Plus). All steers were fed a high-concentrate finishing diet and individual feed intake was recorded daily via the GrowSafe system. BWs were collected every 28 d. On day 55, 10 steers per pen were injected with ovalbumin (OVA). Jugular blood samples were collected from each steer on days 0, 7, 14, and 21 post injection. On day 112, the same steers were injected again with OVA and intramuscularly with a pig red blood cell solution. Jugular blood samples were collected from each steer on days 0, 7, 14, and 21 post injection. On day 124, rumen fluid was collected from 3 steers per treatment and used to estimate in vitro rumen fermentation characteristics. Equal numbers of steers per treatment were transported to a commercial abattoir on days 145, 167, and 185 of the experiment, harvested, and carcass data were collected. Initial BW was similar across treatments. On days 28 and 55, steers receiving DFM had heavier BW (P < 0.01) compared to controls. The average daily gain was greater in DFM-supplemented steers from days 0 to 28 (P < 0.01) and days 0 to 55 (P < 0.01) of the experiment compared to controls. Overall dry matter intake (DMI) was greater (P < 0.04) and overall feed efficiency was similar in DFM-supplemented steers compared to controls. Dressing percentage (P < 0.02) was greater in steers receiving DFM compared to controls. Antibody titers to injected antigens were similar across treatments. However, red blood cell superoxide dismutase activity was greater (P < 0.05) in DFM-supplemented steers compared to controls. In vitro molar proportions of isobutyric and butyric acid were greater (P < 0.01) and dry matter (DM) digestibility tended (P < 0.07) to be greater in rumen fluid obtained from steers supplemented with DFM. These data suggest that BOVAMINE DEFEND Plus supplementation improves growth performance during the initial period of the finishing phase, increases overall DMI and dressing percentage, and may impact antioxidant status in beef cattle.

Design and Synthesis of Novel 3-Nitro-1H-1,2,4-triazole-1,2,3-triazole-1,4-disubstituted Analogs as Promising Antitrypanosomatid Agents: Evaluation of In Vitro Activity against Chagas Disease and Leishmaniasis.

A series of 28 compounds, 3-nitro-1H-1,2,4-triazole, were synthesized by click-chemistry with diverse substitution patterns using medicinal chemistry approaches, such as bioisosterism, Craig-plot, and the Topliss set with excellent yields. Overall, the analogs demonstrated relevant in vitro antitrypanosomatid activity. Analog 15g (R1 = 4-OCF3-Ph, IC50 = 0.09 µM, SI = >555.5) exhibited an outstanding antichagasic activity (Trypanosoma cruzi, Tulahuen LacZ strain) 68-fold more active than benznidazole (BZN, IC50 = 6.15 µM, SI = >8.13) with relevant selectivity index, and suitable LipE = 5.31. 15g was considered an appropriate substrate for the type I nitro reductases (TcNTR I), contributing to a likely potential mechanism of action for antichagasic activity. Finally, 15g showed nonmutagenic potential against Salmonella typhimurium strains (TA98, TA100, and TA102). Therefore, 3-nitro-1H-1,2,4-triazole 15g is a promising antitrypanosomatid candidate for in vivo studies.