Monitoring Coxiella burnetii Infection in Naturally Infected Dairy Sheep Flocks Throughout Four Lambing Seasons and Investigation of Viable Bacteria.

Progression of Coxiella burnetii infection in four naturally infected sheep flocks, and in their farm environment, was monitored throughout four lambing seasons. Flocks with an active infection were selected based on the presence of C. burnetii DNA in bulk-tank milk (BTM) and a high seroprevalence in yearlings during the previous milking period (Spring 2015). During four consecutive lambing seasons (2015/16-2018/19), samples were collected within 1 week after each lambing period from animals (vaginal swabs, milk and feces from ewes, and yearlings) and the environment (dust indoor sheep premises). BTM samples and aerosols (outdoors and indoors) were monthly collected between lambing and the end of milking. Real-time PCR analyses showed different trends in C. burnetii shedding in the flocks, with a general progressive decrease in bacterial shedding throughout the years, interrupted in three flocks by peaks of reinfection associated with specific management practices. A significant relationship was found between C. burnetii fecal shedding and the bacterial burden detected in dust, whereas shedding by vaginal route affected the detection of C. burnetii in indoor aerosols. Three genotypes were identified: SNP8 (three flocks, 52.9% of the samples), SNP1 (two flocks, 44.8% samples), and SNP5 (one flock, two environmental samples). Coxiella burnetii viability in dust measured by culture in Vero cells was demonstrated in two of the flocks, even during the fourth lambing season. The results showed that infection can remain active for over 5 years if effective control and biosafety measures are not correctly implemented.

A Q fever outbreak associated to courier transport of pets.

On August 3rd, 2017, a Q fever outbreak alert was issued at a courier company that in addition to urgent freight transport offered pet delivery services. The epidemiological investigation set the exposition period between June 1 and August 8. In this period, 180 workers from two operational platforms for parcel distribution located in two provinces of the Basque Country (Bizkaia and Araba) were exposed; 64 filled a questionnaire and provided blood samples for serological testing, resulting in 10 confirmed cases (15.6%) and six (9.4%) probable cases. Nine workers (8 confirmed and 1 probable) showed Q fever symptoms, including pneumonia (five cases), and required medical care services, including one hospital admission. The attack rate was 25% (16/64), being higher among workers that visited the Bizkaia platform. This suggested that the origin of the outbreak was in the Bizkaia platform, where animals in transit waited at a pet holding site until being moved to their destination. Environmental samples consisting on 19 surface dust and two aerosol samples were collected at the Bizkaia platform to investigate the presence of C. burnetti DNA. All dust samples were positive by real time PCR, the lowest Ct values being found in dust collected at the pet holding facilities, and therefore suggesting that contamination originated at the pet holding site. The genotype identified in dust was SNP1/MST13, one of the most commonly identified genotypes in goats and sheep in the Basque Country. During the exposure period, two deliveries of miniature goats were made, of which only one could be investigated and tested negative. Although the contamination source could not be unequivocally identified, transport of ruminants was banned at the company, and Q fever was included among the occupational-associated health risks.

Viable Coxiella burnetii in hard cheeses made with unpasteurized milk.

Q fever is a bacterial zoonosis caused by Coxiella burnetii whose main reservoir are small ruminants. Infected animals shed the bacteria into the environment through the products of abortion as well as through feces, urine, and milk. Susceptible people are mainly infected by the inhalation of contaminated aerosols, while food-borne infection is unclear. High prevalence of C. burnetii DNA in cheeses from cattle, sheep or goat has been reported, but studies on viability of C. burnetii in hard cheeses are scarce. In this study, 67 sheep handicraft hard cheeses of different geographic origins made with unpasteurized milk were analyzed for the presence of C. burnetii DNA. To investigate viability of C. burnetii in cheese, 5 cheeses were selected among the 20 that tested DNA positive. Presence of viable C. burnetii was demonstrated in one cheese by experimental inoculation in BALB/c mice and culture in Vero cells. To further investigate the effect of cheese ripening in C. burnetii viability, another 12 cheeses elaborated in the same farm and season, and ripened for between 2.0 and 10.1 months were investigated. Results showed presence of C. burnetii DNA in all of them and viable C. burnetii in 5, indicating that C. burnetii can remain viable after at least 8 months of ripening in hard cheeses made with unpasteurized milk under the acid pH (4.96-5.41) and low water activity (0.9065-0.9533) conditions observed.

Stable levels of Coxiella burnetii prevalence in dairy sheep flocks but changes in genotype distribution after a 10-year period in northern Spain.

Bulk tank milk (BTM) samples were collected from 81 sheep flocks in the Basque Country, Spain, in 2015 and were analysed for antibodies against Coxiella burnetii by ELISA and for C. burnetii DNA by real-time PCR. Thirty-two percent of the flocks had BTM antibodies against C. burnetii. Presence of C. burnetii DNA in BTM was detected in 23% of the flocks, suggesting recent C. burnetii infections. Retrospective data of BTM samples obtained from 154 sheep flocks investigated in 2005 in the same geographic area were compiled to assess temporal changes in C. burnetii infection. The overall percentage of infected sheep flocks did not significantly change after the 10-year period. Among the 46 flocks sampled in both periods, 11 flocks that were negative in 2005 were positive in 2015, 18 maintained their initial status (positive or negative), and 17 positive flocks were negative in 2015. These findings indicate that C. burnetii infection is a dynamic process in dairy sheep in northern Spain. Single nucleotide polymorphism (SNP) genotyping of positive samples identified three genotypes, SNP1 being the most prevalent in 2015 and SNP8 in 2005; SNP4 was only detected once in 2005. These results suggest possible changes in the pattern of genotype infection over time.

A Q Fever Outbreak with a High Rate of Abortions at a Dairy Goat Farm: Coxiella burnetii Shedding, Environmental Contamination, and Viability.

This study describes a Q fever outbreak in a herd of 77 Alpine goats which suffered a high rate of abortions (81% [58/72]) in January 2017 and presents the results of monitoring the contamination and viability of Coxiella burnetii in the farm environment several months after the outbreak. Over the course of 7 months, we studied bacterial shedding by 35 dams with abortions to monitor C. burnetii infection dynamics and the duration of excretion. The highest bacterial shedding load was observed in vaginal mucus, followed by in feces and in milk. Conversely, the duration of C. burnetii shedding was longer through feces (5 months after abortion) than milk (3 months). C. burnetii DNA was detected throughout the study in aerosol samples periodically collected indoors and outdoors from the animal premises. Mouse inoculation and culture in Vero cells demonstrated the presence of viable isolates in dust collected from different surfaces inside the animal facilities during the period of time with the highest number of abortions but not in dust collected 2, 3, and 4 months after the last parturition. Some workers and visitors were affected by Q fever, with attack rates of 78% (7/9) and 31% (4/13), respectively. Affected people mostly showed fever and seroconversion, along with myalgia and arthralgia in two patients and pneumonia in the index case. The genotype identified in animal and environmental samples (SNP1/MST13) turned out to be very aggressive in goats but caused only moderate symptoms in people. After the diagnosis of abortion by Q fever in goats, several control measures were implemented at the farm to prevent contamination inside and outside the animal facilities.IMPORTANCE This work describes a 7-month follow-up of the excretion by different routes of Coxiella burnetii genotype SNP1/MST13 in a herd of goats that suffered high rate of abortions (81%), generating high environmental contamination. Some of the workers and visitors who accessed the farm were infected, with fever as the main symptom but a low incidence of pneumonia. The detected strain (SNP1/MST13 genotype) turned out to be very aggressive in goats. The viability of C. burnetii was demonstrated in the environment of the farm at the time of abortions, but 2 months after the last parturition, no viable bacteria were detected. These results highlighted the importance of implementing good biosafety measures at farms and avoiding the entrance of visitors to farms several months after the end of the kidding period.

Biosynthetic modularity rules in the bisintercalator family of antitumor compounds.

Diverse actinomycetes produce a family of structurally and biosynthetically related non-ribosomal peptide compounds which belong to the chromodepsipeptide family. These compounds act as bisintercalators into the DNA helix. They give rise to antitumor, antiparasitic, antibacterial and antiviral bioactivities. These compounds show a high degree of conserved modularity (chromophores, number and type of amino acids). This modularity and their high sequence similarities at the genetic level imply a common biosynthetic origin for these pathways. Here, we describe insights about rules governing this modular biosynthesis, taking advantage of the fact that nowadays five of these gene clusters have been made public (thiocoraline, triostin, SW-163 and echinomycin/quinomycin). This modularity has potential application for designing and producing novel genetic engineered derivatives, as well as for developing new chemical synthesis strategies. These would facilitate their clinical development.