Fecal microbiota of diarrheic calves: Before, during, and after recovering from disease.

BACKGROUND: It is unknown if gastrointestinal dysbiosis in diarrheic calves causes disease or is a consequence of the disease. OBJECTIVES: Describe the fecal microbiota of calves before, during, and after recovering from diarrhea. ANIMALS: Fifteen female Holstein calves of 0 to 21 days old from a single farm. Seven calves remained healthy throughout the study, and 8 developed diarrhea on Day 14. METHODS: Longitudinal cohort study. Microbiota composition was characterized by amplifying the V4 region of the 16S rRNA gene. RESULTS: Diversity (Shannon index) increased with age in healthy and diarrheic calves from Day 3 to 21, but diarrheic calves had a lower diversity on the day diarrhea was first observed (Day 14). By Day 21, diversity increased in calves that recovered from diarrhea and was not significantly different from that of their healthy counterparts (P > .05). Weighted UniFrac distance showed significant differences in the fecal microbiota between diarrheic and healthy calves at Day 14 of age (PERMANOVA, P < .05), but not before or after diarrhea (PERMANOVA, P > .05). Lactobacillus, Clostridium Sensu Stricto 1, and Collinsella were differentially abundant on Day 10 in calves that developed diarrhea on Day 14 (P < .05). CONCLUSION AND CLINICAL IMPORTANCE: The fecal microbiota of healthy and diarrheic calves evolved similarly during the first 10 days of age but differed significantly on the day of onset of diarrhea. Enriching Lactobacillus, Clostridium Sensu Stricto 1, and Collinsella before diarrhea onset could have been contributed to the development of diarrhea.

Neonatal Calf Diarrhea and Gastrointestinal Microbiota: Etiologic Agents and Microbiota Manipulation for Treatment and Prevention of Diarrhea.

Neonatal calf diarrhea is the leading cause of neonatal morbidity and mortality globally. The changes associated with the gastrointestinal microbiota in neonatal calves experiencing diarrhea and its etiology are not fully understood or completely defined in the literature. Several studies have demonstrated that the fecal microbiota of calves that experience diarrhea substantially deviates from that of healthy age-matched calves. However, one key question remains: whether the changes observed in the bacterial communities (also known as dysbiosis) are a predisposing factor for, or the consequence of, gastrointestinal inflammation caused by the pathogens associated with calf diarrhea. The first objective of this literature review is to present the current information regarding the changes in the fecal microbiota of diarrheic calves and the impact of the pathogens associated with diarrhea on fecal microbiota. Modulation of the gastrointestinal microbiota using pre- and probiotics, colostrum feeding, and fecal microbiota transplantation (FMT) has been used to treat and prevent gastrointestinal diseases in humans and dogs. Although information regarding the use of probiotics for the prevention of diarrhea is available in cattle, little information is available regarding the use of these strategies for treating calf diarrhea and the use of prebiotics or FMT to prevent diarrhea. The second objective of this literature review is to summarize the current knowledge regarding the impact of prebiotics, probiotics, synbiotics, colostrum feeding, and FMT for the treatment and prevention of calf diarrhea.

Nontuberculous Mycobacteria in Horses: A Narrative Review.

Nontuberculous mycobacteria (NTM) infections are increasing in human and veterinary medicine. Although horses were initially thought to be resistant to NTM infection, reports of horses suffering from gastrointestinal, respiratory, and reproductive diseases associated with NTM have increased in the last few decades. The aim of this literature review is to summarize the mycobacteria species found in horses, describe clinical manifestations, diagnostic and treatment approaches, and public health concerns of NTM infection in horses. Clinical manifestations of NTM in horses include pulmonary disease, lymphadenitis, soft tissue, bone infections, and disseminated disease. NTM are also linked to granulomatous enteritis, placentitis, and abortions. Currently, diagnostic methods for NTM are limited and include acid-fast microscopy, bacterial cultures, species-specific PCR assays, and gene sequencing. In humans, NTM treatment guidelines are available, but their application appears inadequate and inconsistent. In horses, treatment guidelines for NTM infections are not available. NTM are a serious public health threat as 70% of people with untreated acquired immunodeficiency syndrome (AIDS) have a chronic pulmonary disease caused by NTM. Thus, it is essential that we gain a better understanding of NTM infections in horses and their zoonotic potential.

Effect of time of sample collection after onset of diarrhea on fecal microbiota composition of calves.

BACKGROUND: The effect of time of sample collection after onset of diarrhea on the fecal microbiota composition of calves is unknown. OBJECTIVE: Compare the fecal microbiota of calves with diarrhea onset on the day of sampling (D <24h), and calves having had diarrhea for >24 to 48 hours (D 24-48h). ANIMALS: Thirty-one diarrheic calves (20 D <24h and 11 D 24-48h), 3 to 7 days of age. METHODS: Cross-sectional study. Diarrhea was defined as a calf with loose feces or watery feces. Assessment of the fecal microbiota was performed by sequencing of 16S ribosomal RNA gene amplicons. RESULTS: Richness and diversity were not statistically different between D <24h and D 24-48h (P > .05), but bacterial membership and structure differed significantly (AMOVA, P < .001 for both comparisons). Linear discriminant analysis effect size (LefSe) showed an enrichment of Faecalibacterium, Phocaeicola, Lachnospiracea, and Lactobacillus in the feces of D <24h calves, whereas Escherichia/Shigella, Ligilactobacillus, Clostridium_Sensu_Stricto, Clostridium_Incerta_Sedis, and Enterococcus were enriched in the D 24-48h calves. CONCLUSION AND CLINICAL IMPORTANCE: Rapid changes in fecal microbiota occur during the first 48 hours of diarrhea with an enrichment of lactic acid-producing bacteria in D <24h followed by an enrichment in Escherichia/Shigella and Clostridium spp. in D 24-48h. The time from diarrhea onset to sampling appears to affect the bacterial composition. Researchers should standardize times for fecal collection based on the time of diarrhea.

Acute diarrhea in horses: A multicenter Canadian retrospective study (2015 to 2019).

This study describes the clinicopathological findings, diagnostic approach, treatment, and factors associated with non-survival of diarrheic horses admitted to 4 Canadian university teaching hospitals between 2015 and 2019. A total of 300 horses, ≥1-year-old, with acute diarrhea were included and represented 1.6% (300/18 481; range: 0.7 to 3%) of admissions during that period, 70% of the horses survived to discharge. Testing for enteropathogens was limited to a single fecal culture for Salmonella spp. in most cases. An enteropathogen was identified in 14% (42/300) of the horses, but in the hospital with higher testing rates enteropathogens were detected in 29% (16/55) of cases. Neorickettsia risticii was the pathogen most frequently detected (31%, 32/102). Antimicrobial drugs and plasma were administered to 57 and 8% of the cases, respectively. Laminitis occurred in 24/298 (8%) of the horses. A multivariable regression model identified an association between non-survival of diarrheic horses and colic signs, increased heart rate, packed cell volume, creatinine concentration, and decreased total protein concentration. A standardized approach for pathogen detection in diarrheic horses is not consistent among Canadian veterinary teaching hospitals, and testing for known pathogens is limited. Signs of colic, severe dehydration, endotoxemia, and hypoproteinemia are associated with non-survival of diarrheic horses.

Diarrhée aiguë chez le cheval : une étude rétrospective canadienne multicentrique (2015 à 2019). Cette étude décrit les résultats clinicopathologiques, l'approche diagnostique, le traitement et les facteurs associés à la non-survie de chevaux diarrhéiques admis dans quatre hôpitaux universitaires canadiens entre 2015 et 2019. Un total de 300 chevaux, ≥1 an, atteints de diarrhée aiguë ont été inclus et représentaient 1,6 % (300/18 481; intervalle : 0,7 à 3 %) des admissions au cours de cette période. Soixante-dix pourcents des chevaux ont survécu jusqu'à leur congé. La recherche d'agents entéropathogènes était limitée à une seule culture fécale pour Salmonella spp. dans la plupart des cas. Un agent entéropathogène a été identifié chez 14 % (42/300) des chevaux, mais à l'hôpital avec des taux de dépistage plus élevés, des agents entéropathogènes ont été détectés dans 29 % (16/55) des cas. Neorickettsia risticii était l'agent pathogène le plus fréquemment détecté (31 %, 32/102). Des médicaments antimicrobiens et du plasma ont été administrés respectivement à 57 et 8 % des cas. Une fourbure est survenue chez 24/298 (8 %) des chevaux. Un modèle de régression multivarié a identifié une association entre la non-survie des chevaux diarrhéiques et les signes de coliques, l'augmentation de la fréquence cardiaque, l'hématocrite, la concentration de créatinine et la diminution de la concentration totale de protéines. Une approche normalisée pour la détection des agents pathogènes chez les chevaux diarrhéiques n'est pas uniforme dans les hôpitaux d'enseignement vétérinaires canadiens, et les tests pour les agents pathogènes connus sont limités. Des signes de coliques, de déshydratation sévère, d'endotoxémie et d'hypoprotéinémie sont associés à la non-survie des chevaux diarrhéiques.(Traduit par Dr Serge Messier).

Calf Diarrhea Is Associated With a Shift From Obligated to Facultative Anaerobes and Expansion of Lactate-Producing Bacteria.

Diarrhea is the leading cause of morbidity, mortality and antimicrobial drug use in calves during the first month of age. Alteration in the bacterial communities of the gastrointestinal tract occurs during diarrhea. Diarrheic calves often develop anion gap (AG) acidosis associated with increased concentrations of unmeasured anions including D- and L-lactate. However, studies investigating the association between gut microbiota alterations and the development of acid-base disorders in diarrheic calves are lacking. We investigated the fecal bacterial alterations of calves with diarrhea and its association with changes in blood pH, and AG. Blood and fecal samples from healthy and diarrheic veal calves were taken 7 days after arrival to the farm. The fecal microbiota of healthy and diarrheic calves was assessed by sequencing of 16S ribosomal RNA gene amplicons. Blood gas analysis was completed using an i-Stat analyzer. In healthy calves, higher richness, evenness, and diversity were observed compared to diarrheic calves. Phocaeicola, Bacteroides, Prevotella, Faecalibacterium, Butyricicoccus, Ruminococcaceae and Lachnospiraceae were enriched in healthy compared with diarrheic calves. Enterococcus, Ligilactobacillus, Lactobacilus, Gallibacterium Streptococcus, and Escherichia/Shigella were enriched in diarrheic calves. In diarrheic calves, an increased abundance of lactate-producing bacteria including Lactobacillus, Streptococcus, Veillonella, Ligilactobacillus and Olsenella was detected. Diarrheic calves had a lower pH and bicarbonate concentration and a higher AG concentration than healthy calves. Together, these results indicate that calf diarrhea is associated with a shift from obligated to facultative anaerobes and expansion of lactate-producing bacteria which are related to acidemia, low bicarbonate and increase AG. Our results highlight the importance of the gastrointestinal microbiota on the clinicopathological changes observed in diarrheic calves.

Hypoxia inducible factor HIF-1 promotes myeloid-derived suppressor cells accumulation through ENTPD2/CD39L1 in hepatocellular carcinoma.

Myeloid-derived suppressor cells (MDSCs) possess immunosuppressive activities, which allow cancers to escape immune surveillance and become non-responsive to immune checkpoints blockade. Here we report hypoxia as a cause of MDSC accumulation. Using hepatocellular carcinoma (HCC) as a cancer model, we show that hypoxia, through stabilization of hypoxia-inducible factor-1 (HIF-1), induces ectoenzyme, ectonucleoside triphosphate diphosphohydrolase 2 (ENTPD2/CD39L1), in cancer cells, causing its overexpression in HCC clinical specimens. Overexpression of ENTPD2 is found as a poor prognostic indicator for HCC. Mechanistically, we demonstrate that ENTPD2 converts extracellular ATP to 5'-AMP, which prevents the differentiation of MDSCs and therefore promotes the maintenance of MDSCs. We further find that ENTPD2 inhibition is able to mitigate cancer growth and enhance the efficiency and efficacy of immune checkpoint inhibitors. Our data suggest that ENTPD2 may be a good prognostic marker and therapeutic target for cancer patients, especially those receiving immune therapy.Myeloid-derived suppressor cells (MDSCs) promote tumor immune escape. Here, the authors show that in hepatocellular carcinoma, hypoxia induces the expression of ENTPD2 on cancer cells leading to elevated extracellular 5'-AMP, which in turn promote the maintenance of MDSCs by preventing their differentiation.

Folate cycle enzyme MTHFD1L confers metabolic advantages in hepatocellular carcinoma.

Cancer cells preferentially utilize glucose and glutamine, which provide macromolecules and antioxidants that sustain rapid cell division. Metabolic reprogramming in cancer drives an increased glycolytic rate that supports maximal production of these nutrients. The folate cycle, through transfer of a carbon unit between tetrahydrofolate and its derivatives in the cytoplasmic and mitochondrial compartments, produces other metabolites that are essential for cell growth, including nucleotides, methionine, and the antioxidant NADPH. Here, using hepatocellular carcinoma (HCC) as a cancer model, we have observed a reduction in growth rate upon withdrawal of folate. We found that an enzyme in the folate cycle, methylenetetrahydrofolate dehydrogenase 1-like (MTHFD1L), plays an essential role in support of cancer growth. We determined that MTHFD1L is transcriptionally activated by NRF2, a master regulator of redox homeostasis. Our observations further suggest that MTHFD1L contributes to the production and accumulation of NADPH to levels that are sufficient to combat oxidative stress in cancer cells. The elevation of oxidative stress through MTHFD1L knockdown or the use of methotrexate, an antifolate drug, sensitizes cancer cells to sorafenib, a targeted therapy for HCC. Taken together, our study identifies MTHFD1L in the folate cycle as an important metabolic pathway in cancer cells with the potential for therapeutic targeting.

Hypoxia induces myeloid-derived suppressor cell recruitment to hepatocellular carcinoma through chemokine (C-C motif) ligand 26.

UNLABELLED: A population of stromal cells, myeloid-derived suppressor cells (MDSCs), is present in tumors. Though studies have gradually revealed the protumorigenic functions of MDSCs, the molecular mechanisms guiding MDSC recruitment remain largely elusive. Hypoxia, O2 deprivation, is an important factor in the tumor microenvironment of solid cancers, whose growth often exceeds the growth of functional blood vessels. Here, using hepatocellular carcinoma as the cancer model, we show that hypoxia is an important driver of MDSC recruitment. We observed that MDSCs preferentially infiltrate into hypoxic regions in human hepatocellular carcinoma tissues and that hypoxia-induced MDSC infiltration is dependent on hypoxia-inducible factors. We further found that hypoxia-inducible factors activate the transcription of chemokine (C-C motif) ligand 26 in cancer cells to recruit chemokine (C-X3-C motif) receptor 1-expressing MDSCs to the primary tumor. Knockdown of chemokine (C-C motif) ligand 26 in cancer cells profoundly reduces MDSC recruitment, angiogenesis, and tumor growth. Therapeutically, blockade of chemokine (C-C motif) ligand 26 production in cancer cells by the hypoxia-inducible factor inhibitor digoxin or blockade of chemokine (C-X3-C motif) receptor 1 in MDSCs by chemokine (C-X3-C motif) receptor 1 neutralizing antibody could substantially suppress MDSC recruitment and tumor growth. CONCLUSION: This study unprecedentedly reveals a novel molecular mechanism by which cancer cells direct MDSC homing to primary tumor and suggests that targeting MDSC recruitment represents an attractive therapeutic approach against solid cancers. (Hepatology 2016;64:797-813).