Shiga Toxin-Producing Escherichia coli (STEC) Associated with Calf Mortality in Uruguay.

In Uruguay, the mortality of dairy calves due to infectious diseases is high. Escherichia coli is a natural inhabitant of the intestinal microbiota, but can cause several infections. The aim of the work was to characterize E. coli isolates from intestinal and extraintestinal origin of dead newborn calves. Using PCR, virulence gene characteristics of pathogenic E. coli were searched. The pathogenic E. coli were molecularly characterized and the phylogroup, serogroup and the Stx subtype were determined. Antibiotic susceptibility was determined using the Kirby-Bauer disk diffusion method and plasmid-mediated quinolone resistance (PMQR) genes with PCR. Finally, clonal relationships were inferred using PFGE. Gene characteristics of the Shiga toxin-producing E. coli (STEC), Enteropathogenic E. coli (EPEC) and Necrotoxigenic E. coli (NTEC) were identified. The prevalence of the iucD, afa8E, f17, papC, stx1, eae and ehxA genes was high and no f5, f41, saa, sfaDE, cdtIV, lt, sta or stx2 were detected. The prevalence of STEC gene stx1 in the dead calves stood out and was higher compared with previous studies conducted in live calves, and STEC LEE+ (Enterohemorrhagic E. coli (EHEC)) isolates with stx1/eae/ehxA genotypes were more frequently identified in the intestinal than in the extraintestinal environment. E. coli isolates were assigned to phylogroups A, B1, D and E, and some belonged to the O111 serogroup. stx1a and stx1c subtypes were determined in STEC. A high prevalence of multi-resistance among STEC and qnrB genes was determined. The PFGE showed a high diversity of pathogenic strains with similar genetic profiles. It can be speculated that EHEC (stx1/eae/ehxA) could play an important role in mortality. The afa8E, f17G1 and papC genes could also have a role in calf mortality. Multidrug resistance defies disease treatment and increases the risk of death, while the potential transmissibility of genes to other species constitutes a threat to public health.

Phenotypic and genotypic survey of antibiotic resistance in Salmonella enterica isolates from dairy farms in Uruguay.

Salmonella enterica is an important zoonotic pathogen that is frequently identified in dairy farming systems. An increase in antibiotic resistance has led to inadequate results of treatments, with impacts on animal and human health. Here, the phenotypic and genotypic susceptibility patterns of Salmonella isolates from dairy cattle and dairy farm environments were evaluated and compared. A collection of 75 S. enterica isolates were evaluated, and their phenotypic susceptibility was determined. For genotypic characterization, the whole genomes of the isolates were sequenced, and geno-serotypes, sequence types (STs) and core-genome-sequence types were determined using the EnteroBase pipeline. To characterize antibiotic resistance genes and gene mutations, tools from the Center for Genomic Epidemiology were used. Salmonella Dublin (SDu), S. Typhimurium (STy), S. Anatum (SAn), S. Newport (SNe), S. Agona (Sag), S. Montevideo (SMo) and IIIb 61:i:z53 were included in the collection. A single sequence type was detected per serovar. Phenotypic non-susceptibility to streptomycin and tetracycline was very frequent in the collection, and high non-susceptibility to ciprofloxacin was also observed. Multidrug resistance (MDR) was observed in 42 isolates (56.0%), with SAn and STy presenting higher MDR than the other serovars, showing non-susceptibility to up to 6 groups of antibiotics. Genomic analysis revealed the presence of 21 genes associated with antimicrobial resistance (AMR) in Salmonella isolates. More than 60% of the isolates carried some gene associated with resistance to aminoglycosides and tetracyclines. Only one gene associated with beta-lactam resistance was found, in seven isolates. Two different mutations were identified, parC_T57S and acrB_R717Q, which confer resistance to quinolones and azithromycin, respectively. The accuracy of predicting antimicrobial resistance phenotypes based on AMR genotypes was 83.7%. The genomic approach does not replace the phenotypic assay but offers valuable information for the survey of circulating antimicrobial resistance. This work represents one of the first studies evaluating phenotypic and genotypic AMR in Salmonella from dairy cattle in South America.

Diagnostic Investigation of 100 Cases of Abortion in Sheep in Uruguay: 2015-2021.

The aim of this work was to identify causes of abortion through laboratory investigations in sheep flocks in Uruguay. One hundred cases of abortion, comprising 58 fetuses, 36 fetuses with their placentas, and 6 placentas were investigated in 2015-2021. Cases were subjected to gross and microscopic pathologic examinations, and microbiological and serological testing for the identification of causes of abortion, including protozoal, bacterial, and viral pathogens. An etiologic diagnosis was determined in 46 (46%) cases, including 33 (33%) cases caused by infectious pathogens, as determined by the detection of a pathogen along with the identification of fetoplacental lesions attributable to the detected pathogen. Twenty-seven cases (27%) were caused by Toxoplasma gondii, 5 (5%) by Campylobacter fetus subspecies fetus, and 1 (1%) by an unidentified species of Campylobacter. Fourteen cases (14%) had inflammatory and/or necrotizing fetoplacental lesions compatible with an infectious etiology. Although the cause for these lesions was not clearly identified, T. gondii was detected in 4 of these cases, opportunistic bacteria (Bacillus licheniformis, Streptococcus sp.) were isolated in 2 cases, and bovine viral diarrhea virus 1 subtype i (BVDV-1i) was detected in another. Campylobacter jejuni was identified in 1 (1%) severely autolyzed, mummified fetus. BVDV-2b was identified incidentally in one fetus with an etiologic diagnosis of toxoplasmosis. Microscopic agglutination test revealed antibodies against ≥1 Leptospira serovars in 15/63 (23.8%) fetuses; however, Leptospira was not identified by a combination of qPCR, culture, fluorescent antibody testing nor immunohistochemistry. Neospora caninum, Chlamydia abortus, Chlamydia pecorum, Coxiella burnetii and border disease virus were not detected in any of the analyzed cases. Death was attributed to dystocia in 13 (13%) fetuses delivered by 8 sheep, mostly from one highly prolific flock. Congenital malformations including inferior prognathism, a focal hepatic cyst, and enterohepatic agenesis were identified in one fetus each, the latter being the only one considered incompatible with postnatal life. Toxoplasmosis, campylobacteriosis and dystocia were the main identified causes of fetal losses. Despite the relatively low overall success rate in establishing an etiologic diagnosis, a systematic laboratory workup in cases of abortion is of value to identify their causes and enables zoonotic pathogens surveillance.

Fiebre Q: revisión histórica de casos humanos en Uruguay. Abordaje desde la complementariedad entre las ciencias médicas y veterinarias / Q fever: historical review of human cases in Uruguay. A complementary approach from the medical and veterinarian sciences / Febre Q: revisão histórica de casos humanos no Uruguai. Abordagem a partir da complementaridade entre as ciências médicas e veterinárias

Resumen: La fiebre Q es una zoonosis distribuida mundialmente, causada por Coxiella burnetii. Los bovinos, ovinos y caprinos son la fuente más frecuente de infección en humanos, en los que la enfermedad es de notificación obligatoria ante el Ministerio de Salud Pública. Revisamos las publicaciones que describen casos de fiebre Q en humanos en Uruguay, con foco en sus características epidemiológicas, y discutimos las pruebas diagnósticas disponibles localmente. Se incluyeron nueve trabajos publicados e información del registro de enfermedades profesionales. Colectivamente fueron analizadas 2.715 personas con sospecha de fiebre Q entre 1956-2019, siendo 959 (35,3%) seropositivas. Los diagnósticos se basaron en serología, clínica y/o antecedentes de exposición laboral. Epidemiológicamente, el ganado o material proveniente del mismo fueron considerados las fuentes más probables de exposición en la mayoría de los casos. Según el ámbito de ocurrencia, no sistemáticamente reportado, los casos se registraron principalmente por exposición a ovinos y bovinos en frigoríficos o en la cadena cárnica (positivos/evaluados: 863/1540; seropositivos sintomáticos: 585); y en menor medida a bovinos lecheros (sector lácteo (positivos/evaluados:20/58; seropositivos sintomáticos: 17) y laboratorio diagnóstico (positivos/evaluados:2/4; seropositivos sintomáticos: 1)), o rumiantes silvestres (seropositivos/evaluados:25/117; seropositivos sintomáticos: 17). Hipertermia, cefaleas y sudoración fueron reportados. La inhalatoria fue asumida como la vía de infección en todos los casos. Actualmente no están disponibles localmente pruebas de PCR para detección de C. burnetii en humanos, siendo una limitante en el diagnóstico, particularmente en etapas tempranas. La colaboración interdisciplinaria de profesionales de salud humana y animal es esencial en el abordaje de esta zoonosis.

Summary: Q fever is a globally distributed zoonosis caused by the bacteria Coxiella burnetii. Bovines, sheep and goats are the most frequent source of infection in humans, and it is mandatory for the latter to report the disease to the Ministry of Public Health. We reviewed the literature describing cases of Q fever in humans in Uruguay, focusing on epidemiological characteristics, and we discussed the diagnostic tests locally available. Nine published studies were included in the review, as well as the information in the professional diseases registry. 2.715 people with a suspicion of Q fever were collectively analyzed between 1956-2019, 959 (55.3%) of them being seropositive. Diagnosis were based on serology, clinical examination and/or a history of exposure while working. Epidemiologically, the cattle or material originating in it were considered as the most probable sources of exposure in most cases. Depending on the context cases arose, which were not systematically reported, they were mainly caused by exposure to sheep and cows in meat processing plants or in the meat chain (positive/evaluated: 863/1540; seropositive or symptomatic: 585); and to a lesser extent dairy cattle (milk sector (positive/evaluated: 20/58; symptomatic seropositive: 17) and diagnostic laboratory (positive/evaluated: 2/4; symptomatic seropositive: 1)) or wild ruminants (positive/evaluated: 25/117; symptomatic seropositive: 17). Hyperthermia, headaches and sweating were reported. In all cases inhalation was adopted as the source of infection. Today, there are no PCR tests to detect C. burnetii in humans available locally, what constitutes a limitation to diagnosis, in particular in early stages. Interdisciplinary collaboration between animal and human health professionals is key to approach this zoonosis.

Resumo: A febre Q é uma zoonose mundial causada por Coxiella burnetii. Bovinos, ovinos e caprinos são a fonte mais frequente de infecção em humanos, nos quais a doença é de notificação compulsória ao Ministério da Saúde Pública. Revisamos as publicações que descrevem casos de febre Q em humanos no Uruguai, enfocando em suas características epidemiológicas, e discutimos os testes diagnósticos disponíveis localmente. Foram incluídos nove trabalhos publicados e informações do registro de doenças ocupacionais. Foram analisadas 2.715 pessoas com suspeita de febre Q entre 1956-2019, das quais 959 (35,3%) eram soropositivas. Os diagnósticos foram baseados na sorologia, sintomas e/ou história de exposição ocupacional. Do ponto de vista epidemiológico, o gado ou material preveniente de gado foram considerados as fontes mais prováveis de exposição na maioria dos casos. De acordo com o lugar de ocorrência, não notificado sistematicamente, os casos foram registrados principalmente por exposição a ovinos e bovinos em matadouros ou na cadeia da carne (positivos/avaliados: 863/1540; soropositivos sintomáticos: 585); e em menor proporção gado leiteiro (setor leiteiro (positivo/avaliado: 20/58; soropositivo sintomático: 17) e laboratório de diagnóstico (positivo/avaliado: 2/4; soropositivo sintomático: 1)), ou ruminantes silvestres (soropositivo/avaliado: 25/117; soropositivos sintomáticos: 17). Hipertermia, dores de cabeça e sudorese foram relatados. A inalação foi assumida como via de infecção em todos os casos. Atualmente, os testes de PCR para detecção de C. burnetii em humanos não estão disponíveis localmente, sendo uma limitação no diagnóstico, principalmente em estágios iniciais. A colaboração interdisciplinar de profissionais de saúde humana e animal é essencial no enfrentamento dessa zoonose.

Encefalitis asociada a astrovirus bovino neurotrópico, ¿una enfermedad subdiagnosticada en Sudamérica? / Neurotropic bovine astrovirus-associated encephalitis: An underdiagnosed diseasein South America?

Resumen Describimos un caso de encefalitis asociada a infección por astrovirus bovino neu-rotrópico en una vaca lechera, raza Jersey, del departamento de San José, Uruguay. Esterepresenta el segundo caso reportado de esta condición en el hemisferio sur. La vaca, únicaafectada de un rodeo de 70 bovinos, manifestó signos clínicos neurológicos con curso de 2 días,luego de los que murió espontáneamente. El examen histopatológico reveló meningoencefalitislinfocítica, histiocítica y plasmacítica, con necrosis neuronal, sin cuerpos de inclusión. No sedetectaron en el cerebro otros agentes infecciosos, incluyendo el virus de la rabia (Lyssavirus),alfaherpesvirus bovino-1 y alfaherpesvirus bovino-5 (Varicellovirus), virus de la diarrea viralbovina (Pestivirus), virus del Nilo Occidental (Flavivirus), Listeria monocytogenes, Histophi-lus somni y otras bacterias. Dado que el descubrimiento de astrovirus neurotrópicos en variasespecies de mamíferos, incluidos humanos, es reciente, proponemos que los casos de encefalitis por astrovirus pudieron haber pasado inadvertidos en Sudamérica. Discutimos brevementeel diagnóstico patológico diferencial de encefalitis infecciosas en bovinos.

Abstract We describe a case of neurotropic bovine astrovirus-associated encephalitis in a Jer-sey dairy cow from the department of San José, Uruguay. This represents the second case of thiscondition reported in the Southern Hemisphere. The cow was the only one affected in a herd of70 cows, showing neurological signs with a 2-day clinical course, before dying spontaneously.Histopathological examination revealed lymphocytic, histiocytic, and plasmacytic meningoen-cephalitis with neuronal necrosis, without detectable inclusion bodies. Other infectious agents,including Rabies virus (Lyssavirus), Bovine alphaherpesvirus-1 and Bovine alphaherpesvirus-5(Varicellovirus), Bovine viral diarrhea virus (Pestivirus), West Nile virus (Flavivirus), Listeriamonocytogenes, Histophilus somni and other bacteria, were not detected in the brain. We pro-pose that given the recent discovery of neurotropic astroviruses in various mammalian species,including humans, cases of astrovirus encephalitis may have gone undetected in South America.We briefly discuss the differential pathologic diagnosis of infectious bovine encephalitis.

Placentitis and abortion caused by a multidrug resistant strain of Campylobacter fetus subspecies fetus in a sheep in Uruguay / Placentitis y aborto causado por una cepa de Campylobacter fetus subespecie fetus multirresistente a antibióticos en una oveja en Uruguay

Abstract Campylobacter fetus fetus (Cff) is a major infectious cause of abortion in sheep worldwide, and an opportunistic human pathogen. Information on Cff as an ovine abortifacient in South America is limited. We describe a case of abortion caused by a multidrug resistant strain of Cff in a sheep in Uruguay. In August 2017, 3/57 pregnant ewes (5.3%) aborted whithin one week. Histopathologic examination of the placenta of an aborted ewe revealed severe neutrophilic and fibrinonecrotizing placentitis with vasculitis and thrombosis of the chorionic arterioles. Cff was isolated on microaerobic culture in Skirrow agar, and further confirmed by 16S rDNA PCR amplification and sequencing, and endpoint and real time PCR assays. Antimicrobial sensitivity testing revealed resistance to tetracyclines, nalidixic acid, telithromycin and clindamycin. Other abortifacients were not detected. Further studies are necessary to determine the geographic distribution, ecology, epidemiology, economic impact, and antimicrobial resistance of Cff in sheep flocks in Uruguay.

Resumen Campylobacter fetus fetus (Cff) es una importante causa de abortos en ovinos y un patógeno oportunista en humanos. La información sobre Cff como abortifaciente en ovinos en Sudamérica es limitada. Describimos un caso de aborto causado por una cepa de Cff mul tirresistente a antibióticos en una oveja en Uruguay. En agosto de 2017, 3/57 ovejas prenadas (5,3%) abortaron en una semana. El examen histopatológico de la placenta de una de ellas reveló placentitis neutrofílica fibrinonecrosante severa, vasculitis y trombosis. Cff fue aislado en microaerobiosis en agar Skirrow, y confirmado mediante amplificación del ADNr 16S por PCR seguida de secuenciación, y por PCR punto final y qPCR. Las pruebas de sensibilidad antimicrobiana revelaron resistencia a tetraciclinas, ácido nalidíxico, telitromicina y clindamicina. No se detectaron otros abortifacientes. Son necesarios más estudios para determinar la distribución geográfica, ecología, epidemiología, el impacto económico y la resistencia antimicrobiana de Cff en majadas ovinas de Uruguay.

Placentitis and abortion caused by a multidrug resistant strain of Campylobacter fetus subspecies fetus in a sheep in Uruguay.

Campylobacter fetusfetus (Cff) is a major infectious cause of abortion in sheep worldwide, and an opportunistic human pathogen. Information on Cff as an ovine abortifacient in South America is limited. We describe a case of abortion caused by a multidrug resistant strain of Cff in a sheep in Uruguay. In August 2017, 3/57 pregnant ewes (5.3%) aborted whithin one week. Histopathologic examination of the placenta of an aborted ewe revealed severe neutrophilic and fibrinonecrotizing placentitis with vasculitis and thrombosis of the chorionic arterioles. Cff was isolated on microaerobic culture in Skirrow agar, and further confirmed by 16S rDNA PCR amplification and sequencing, and endpoint and real time PCR assays. Antimicrobial sensitivity testing revealed resistance to tetracyclines, nalidixic acid, telithromycin and clindamycin. Other abortifacients were not detected. Further studies are necessary to determine the geographic distribution, ecology, epidemiology, economic impact, and antimicrobial resistance of Cff in sheep flocks in Uruguay.

Probiotic Lactobacilli Administration Induces Changes in the Fecal Microbiota of Preweaned Dairy Calves.

Early microbial colonization is a determinant factor in animal health, and probiotic administration has been demonstrated to modulate intestinal microbiota and promote health in dairy calves. The objective of this study was to evaluate changes in calves' fecal microbiota after the administration of two probiotic lactobacilli strains that had previously exhibited beneficial effects in calves' health in relation to neonatal calf diarrhea. An in vivo assay was performed with 30 newborn male Holstein calves that were divided into three groups. Two groups were orally administered with two different lactobacilli strains (Lactobacillus johnsonii TP1.6 or Limosilactobacillus reuteri TP1.3B), and the third was the control group. Calves (5 to 9 days old) were administered with freeze-dried bacteria once a day for 10 consecutive days. Feces samples were taken before the first administration (day 0) and then again on days 10 and 21, and the V4 region of the bacterial 16S ribosomal gene was sequenced with an Illumina MiSeq 250 paired-end platform. The administration of both strains significantly affected the total bacterial community composition, and the effect lasted for 11 days after the last dose. In particular, amplicon sequence variants related to Bifidobacterium and Akkermansia genera were significantly higher in both treated groups. Therefore, modulation of the intestinal microbiota is a potential mechanism of action behind the beneficial effects of these probiotic strains.

[Neurotropic bovine astrovirus-associated encephalitis: An underdiagnosed disease in South America?] / Encefalitis asociada a astrovirus bovino neurotrópico, ¿una enfermedad subdiagnosticada en Sudamérica?

We describe a case of neurotropic bovine astrovirus-associated encephalitis in a Jersey dairy cow from the department of San José, Uruguay. This represents the second case of this condition reported in the Southern Hemisphere. The cow was the only one affected in a herd of 70 cows, showing neurological signs with a 2-day clinical course, before dying spontaneously. Histopathological examination revealed lymphocytic, histiocytic, and plasmacytic meningoencephalitis with neuronal necrosis, without detectable inclusion bodies. Other infectious agents, including Rabies virus(Lyssavirus), Bovine alphaherpesvirus-1 and Bovine alphaherpesvirus-5(Varicellovirus), Bovine viral diarrhea virus(Pestivirus), West Nile virus(Flavivirus), Listeria monocytogenes, Histophilus somni and other bacteria, were not detected in the brain. We propose that given the recent discovery of neurotropic astroviruses in various mammalian species, including humans, cases of astrovirus encephalitis may have gone undetected in South America. We briefly discuss the differential pathologic diagnosis of infectious bovine encephalitis.

Serological Evidence of Human Infection with Coxiella burnetii after Occupational Exposure to Aborting Cattle.

Cattle are broadly deemed a source of Coxiella burnetii; however, evidence reinforcing their role in human infection is scarce. Most published human Q fever outbreaks relate to exposure to small ruminants, notably goats. Anti-phase II C. burnetii IgG and IgM were measured by indirect fluorescent antibody tests in 27 farm and veterinary diagnostic laboratory workers to ascertain whether occupational exposure to cattle aborting due to C. burnetii was the probable source of exposure. Four serological profiles were identified on the basis of anti-phase II IgG and IgM titres. Profile 1, characterised by high IgM levels and concurrent, lower IgG titres (3/27; 11.1%); Profile 2, with both isotypes with IgG titres higher than IgM (2/27; 7.4%); Profile 3 with only IgG phase II (5/27; 18.5%); and Profile 4, in which neither IgM nor IgG were detected (17/27; 63.0%). Profiles 1 and 2 are suggestive of recent C. burnetii exposure, most likely 2.5-4.5 months before testing and, hence, during the window of exposure to the bovine abortions. Profile 3 suggested C. burnetii exposure that most likely predated the window of exposure to aborting cattle, while Profile 4 represented seronegative individuals and, hence, likely uninfected. This study formally linked human Q fever to exposure to C. burnetii infected cattle as a specific occupational hazard for farm and laboratory workers handling bovine aborted material.

Coxiella burnetii abortion in a dairy farm selling artisanal cheese directly to consumers and review of Q fever as a bovine abortifacient in South America and a human milk-borne disease.

Coxiella burnetii is a highly transmissible intracellular bacterium with a low infective dose that causes Q fever (coxiellosis), a notifiable zoonotic disease distributed worldwide. Livestock are the main source of C. burnetii transmission to humans, which occurs mostly through the aerogenous route. Although C. burnetii is a major abortifacient in small ruminants, it is less frequently diagnosed in aborting cattle. We report a case of C. burnetii abortion in a lactating Holstein cow from a dairy farm producing and selling artisanal cheese directly to consumers in Uruguay, and review the literature on coxiellosis as a bovine abortifacient in South America and as a milk-borne disease. The aborted cow had severe necrotizing placentitis with abundant intratrophoblastic and intralesional C. burnetii confirmed by immunohistochemistry and PCR. After primo-infection in cattle, C. burnetii remains latent in the lymph nodes and mammary glands, with milk being a significant and persistent excretion route. Viable C. burnetii has been found in unpasteurized milk and cheeses after several months of maturing. The risk of coxiellosis after the consumption of unpasteurized dairy products, including cheese, is not negligible. This report raises awareness on bovine coxiellosis as a potential food safety problem in on-farm raw cheese manufacturing and sales. The scant publications on abortive coxiellosis in cattle in South America suggest that the condition has probably gone underreported in all countries of this subcontinent except for Uruguay. Therefore, we also discuss the diagnostic criteria for laboratory-based confirmation of C. burnetii abortion in ruminants as a guideline for veterinary diagnosticians.

Biogenic Silver Nanoparticles as a Strategy in the Fight Against Multi-Resistant Salmonella enterica Isolated From Dairy Calves.

Infectious diseases are one of the most important health problems worldwide, one of the main causes being the development of multi-resistant microorganisms. Likewise, the zoonotic potential of some pathogens and their ability to transfer resistance mechanisms, reduce the therapeutic options in both humans and animals. Salmonella enterica is an important pathogen that affects a wide range of animal species and humans, being Salmonella Typhimurium one of the most frequent serotypes affecting cattle, causing enteritis, diarrhea, and septicemia. The search for alternative therapeutic approaches has gained importance since the emergence of multidrug resistance to antibiotics and periodic outbreaks of salmonellosis. In this sense, the discovery of new drugs and the development of new strategies, such as the use of nanoparticles with antimicrobial activity, are very promising. The aim of this work was the extracellular production of biogenic silver nanoparticles using fungal extracts and the evaluation of their antimicrobial activity against resistant and multi-resistant Salmonella Typhimurium strains. We here demonstrated the potential of the biogenic nanoparticles as effective bacteriostatic and bactericidal agents for use in biomedical applications. In addition, Confocal Raman Microscopy and Atomic Force Microscopy were used to advance the understanding of the antimicrobial mechanism of biogenic nanoparticles against these pathogenic strains, the results of which suggested that the nanoparticles produced damage in several bacterial cell structures.

Molecular prevalence of Coxiella burnetii in bulk-tank milk from bovine dairy herds: Systematic review and meta-analysis.

Coxiella burnetii is an obligate intracellular zoonotic bacterium that causes Q fever. Ruminants, including cattle, are broadly known to be reservoirs for this bacterium. Since 2006, many research groups have evaluated the herd-level prevalence of C. burnetii in cattle by molecular techniques on composite milk samples. This study explored the global C. burnetii herd-level prevalence from studies done on bovine bulk-tank milk (BTM) samples using PCR-based analysis. Also, moderators were investigated to identify sources of heterogeneity. Databases (CAB Abstracts, Medline via Ovid, PubMed, Web of Science and Google Scholar) were searched for index articles on C. burnetii prevalence in BTM samples by PCR published between January-1973 and November-2018. Numerous studies (1054) were initially identified, from which seventeen original publications were included in the meta-analysis based on the pre-defined selection criteria. These studies comprised 4031 BTM samples from twelve countries. A random-effects model was used because of considerable heterogeneity (I 2 = 98%) to estimate the herd-level prevalence of C. burnetii as 37.0%(CI95%25.2-49.5%). The average herd size appeared to account for a high level of the heterogeneity. No other moderators (geographic location, gross national income or notification criteria for Q fever) seemed to be determinant. This systematic evaluation demonstrated a high molecular prevalence of C. burnetii in BTM samples both in European and non-European countries, evidencing a widespread herd-level circulation of this agent in bovine dairy farms around the world. Meta-regression showed herd size as the most relevant moderator with the odds of a BTM sample testing positive doubling with every unit increase.

Causes of neonatal calf diarrhea and mortality in pasture-based dairy herds in Uruguay: a farm-matched case-control study.

Neonatal calf diarrhea (NCD) and mortality cause significant losses to the dairy industry. The preweaning dairy calf mortality risk in Uruguay is high (15.2%); however, causes for these losses are largely unknown. This study aimed to assess whether various pathogens were associated with NCD and death in Uruguayan dairy calves and whether these infections, diarrhea, or deaths were associated with the failure of transfer of passive immunity (FTPI). Contemporary diarrheic (n = 264,) and non-diarrheic (n = 271) 1- to 30-day-old calves from 27 farms were sampled. Feces were analyzed by antigen-capture ELISA for Cryptosporidium spp., rotavirus, bovine coronavirus, and Escherichia coli F5+, RT-PCR for bovine astrovirus (BoAstV), and bacterial cultures for Salmonella enterica. Blood/serum was analyzed by RT-PCR or antigen-capture ELISA for bovine viral diarrhea virus (BVDV). Serum of ≤ 8-day-old calves (n = 95) was assessed by refractometry to determine the concention of serum total proteins (STP) as an indicator of FTPI. Whether the sampled calves died before weaning was recorded. At least one pathogen was detected in 65.4% of the calves, and this percentage was significantly higher in diarrheic (83.7%) versus non-diarrheic (47.6%) calves. Unlike the other pathogens, Cryptosporidium spp. and rotavirus were associated with NCD. Diarrheic calves, calves infected with any of the pathogens, and calves infected with rotavirus had significantly lower concentrations of STP. Diarrheic calves had higher chances of dying before weaning than non-diarrheic calves. Diarrheic calves infected with S. enterica were at increased risk of mortality. Controlling NCD, salmonellosis, cryptosporidiosis, and rotavirus infections, and improving colostrum management practices would help to reduce calf morbi-mortality in dairy farms in Uruguay.

Accurate and fast identification of Campylobacter fetus in bulls by real-time PCR targeting a 16S rRNA gene sequence.

Campylobacter fetus is an important animal pathogen that causes infectious infertility, embryonic mortality and abortions in cattle and sheep flocks. There are two recognized subspecies related with reproductive disorders in livestock: Campylobacter fetus subsp. fetus (Cff) and Campylobacter fetus subsp. venerealis (Cfv). Rapid and reliable detection of this pathogenic species in bulls is of upmost importance for disease control in dairy and beef herds as they are asymptomatic carriers. The aim of the present work was to assess the performance a real-time PCR (qPCR) method for the diagnosis of Campylobacter fetus in samples from bulls, comparing it with culture and isolation methods. 520 preputial samples were both cultured in Skirrow's medium and analyzed by qPCR. The estimated sensitivity of qPCR was 90.9% (95% CI, 69.4%-100%), and the specificity was 99.4% (95% CI, 98.6% - 100%). The proportion of C. fetus positive individuals was 2.1% by isolation and 2.5% by qPCR. Isolates were identified by biochemical tests as Cfv (n = 9) and Cff (n = 2). Our findings support the use of qPCR for fast and accurate detection of C. fetus directly from field samples of preputial smegma of bulls. The qPCR method showed to be suitable for massive screenings because it can be performed in pooled samples without losing accuracy and sensitivity.

Virulence genes of Escherichia coli in diarrheic and healthy calves.

Escherichia coli ETEC, EPEC, NTEC and STEC/EHEC pathotypes are often isolated from bovine feces. The objective of this study was to detect 21 E. coli virulence genes in feces from 252 dairy calves in Uruguay (149 with neonatal diarrhea - NCD - and 103 asymptomatic). Genes iucD, f17A, afa8E, papC, clpG and f17G(II) were the most prevalent (81.3%; 48.4%; 37.3%; 35.7%; 34.1%; 31.3%, respectively). Genes eae, stx1and stx2 were poorly represented; 13/252 animals harbored one or a combination of these genes. The prevalence of the cnf gene was 4.4%, while that of cdt-IV and cdt-III genes was 24.2% and 12.7% respectively. This study reports updated data about the virulence profiles of E. coli in dairy calves in Uruguay. A large number of adhesins and toxin genes were detected. Our results demonstrate that E. coli from bovine feces has diarrheagenic and extraintestinal profiles although other NCD risks factors may contribute to the disease outcome.

Transferable Resistance to Highest Priority Critically Important Antibiotics for Human Health in Escherichia coli Strains Obtained From Livestock Feces in Uruguay.

The aim of this work was to detect Escherichia coli isolates displaying resistance to oxyimino-cephalosporins, quinolones, and colistin in feces from livestock in Uruguay. During 2016-2019, fecal samples from 132 broiler and layer chicken flocks, 100 calves, and 50 pigs, were studied in Uruguay. Samples were cultured on MacConkey Agar plates supplemented with ciprofloxacin, ceftriaxone, or colistin. E. coli isolates were identified by mass spectrometry and antibiotic susceptibility testing was performed by disk diffusion agar method and colistin agar test. Antibiotic resistance genes were detected by polymerase chain reaction and sequencing. The most frequently detected resistance gene was qnrB19, recovered from 87 animals. Regarding plasmid-mediated quinolone resistance genes, qnrS1 was the second in prevalence (23 animals) followed by qnrE1, found in 6 chickens and two calves. Regarding resistance to oxyimino-cephalosporins, 8 different ß-lactamase genes were detected: bla CTX-M-8 and bla CMY-2 were found in 23 and 19 animals, respectively; next, bla CTX-M-2 and bla SHV-12 in 7 animals each, followed by bla CTX-M-14 in 5, bla CTX-M-15 and bla SHV2a in 2, and bla CTX-M-55 in a single animal. Finally, the mcr-1 gene was detected only in 8 pigs from a single farm, and in a chicken. Isolates carrying bla CMY-2 and bla SHV-12 were also found in these animals, including two isolates featuring the bla CMY-2/mcr-1 genotype. To the best of our knowledge, this is the first work in which the search for transferable resistance to highest priority critically important antibiotics for human health is carried out in chickens and pigs chains of production animals in Uruguay.

Complete Genome Sequence of Campylobacter fetus Isolated from a Sheep.

Campylobacter fetus is an important reproductive pathogen of ruminants that occasionally infects humans. Here, we describe the complete circularized genome of a strain of Campylobacter fetus subsp. fetus isolated from a sheep. The final assembly consisted of a unique contig with a length of 1,849,237 bp.