Dyego Gonçalves Lino Borges

Ivermectin and moxidectin resistance characterization by larval migration inhibition test in field isolates of Cooperia spp. in beef cattle, Mato Grosso do Sul, Brazil.

Ivermectin (IVM) resistance of Cooperia spp. in cattle has become an increasing and global problem. The early detection of anthelmintic resistance (AR) is important to propose strategies to slow down the development of resistance and requires sensitive, reliable, economic high-throughput and practical tests. The purpose of the present study was to apply a larval migration inhibition test (LMIT) for evaluating IVM and MOX efficacy against well-characterized field isolates of Cooperia spp. infecting cattle in Brazil. Eight isolates were used for IVM and seven for MOX. The following EC50 values of IVM were observed for the isolates: susceptible, 1.16 ηmol; Nova Alvorada do Sul I, 4.09 ηmol (RF=3.52); Campo Grande BNA, 3.57 ηmol (RF=3.07); Campo Grande TBR, 4.09 ηmol (RF=3,52); Nova Alvorada do Sul II, 2.50 ηmol (RF=2.15); Bandeirantes, 11.35 ηmol (RF=9.78); Campo Grande II, 6.03 ηmol (RF=5.20); and Porto Mortinho, 8.63 ηmol (RF=7.44). For MOX, the following EC50 values were observed: susceptible, 0.75 ηmol; Campo Grande BNA, 0.93 ηmol (RF=1.24); Campo Grande TBR, 0.36 ηmol (RF=0.48); Nova Alvorada do Sul II, 2.57 ηmol (RF=3.42); Bandeirantes, 1.43 ηmol (RF=1.90); Campo Grande II, 1.08 ηmol (RF=1.44); and Porto Mortinho, 0.49 ηmol (RF=0.65). The LMIT used in the present study can be a useful tool for in vitro evaluation of IVM, but not of MOX. However, such methodology cannot be used in large-scale studies yet. The isolates of Cooperia spp. showed various degrees of resistance to IVM, though remaining susceptible to MOX.

P-gp modulating drugs greatly potentiate the in vitro effect of ivermectin against resistant larvae of Haemonchus placei.

Since its production in the 1980s, ivermectin (IVM) has been used indiscriminately and the selection pressure to which bovine gastrointestinal nematodes have been exposed has been intense, resulting in considerable economic losses due to parasitic resistance. One possibility for the control of resistant parasites is the use of P-glycoprotein (P-gp) modulators, because one of the main biochemical changes in ivermectin-resistant parasites is the increased activity of membrane proteins responsible for the efflux of drugs and xenobiotics. This study aimed to evaluate the in vitro effect of eight P-gp modulating drugs to potentiate IVM efficacy against an IVM-resistant field isolate of Haemonchus placei (Nematoda: Trichostrongylidae). The association of IVM with cyclosporin-A, ceftriaxone, dexamethasone, diminazene aceturate, quercetin, trifluoperazine, verapamil, or vinblastine resulted in increased IVM (10(-4)M) efficacy of 5.1%, 49.06%, 76.42%, 3.31%, 28.85%, 13.74%, 45.64% and 43.61%, respectively, and reduced the IVM half maximal effective concentration (EC50) from 4.381 × 10(-6)M to 9.877 × 10(-8), 2.739 × 10(-7), 1.240 × 10(-6), 1.651 × 10(-6), 2.710 × 10(-7), 1.159 × 10(-7), 1.026 × 10(-6) and 7.136 × 10(-7)M, respectively. Only diminazene aceturate did not significantly reduce the number of migrating larvae when associated with IVM (P > 0.05). The effect of P-gp modulating drugs depended on IVM concentration, with greater potentiating effect at lower IVM concentrations. The in vitro application of trifluoperazine, dexamethasone, quercetin, verapamil, cyclosporin A, vinblastine, and ceftriaxone potentiated IVM efficacy against an IVM-resistant field isolate of H. placei, resulting in higher efficacy and lower IVM EC50.

Efficacy of extracts from plants of the Brazilian Pantanal against Rhipicephalus (Boophilus) microplus

This research evaluated the in vitro acaricidal activity of extracts from 21 plant species from the Pantanal of Mato Grosso do Sul. During stage I, a larval immersion test was performed using three extract concentrations (5%, 20%, and 40%). During stage II, we used only plants that showed over 95% efficiency at the 40% concentration in stage I in an amount sufficient for the adult immersion test. Aeschynomene denticulata, Angelonia hirta, Aspilia latissima, Caperonia castaneifolia, Centratherum punctatum, Crotalaria micans, Diodia kuntzei, Echinodorus paniculatus, Hyptis mutabilis, Lantana canescens, Melanthera latifolia, Ocotea diospyrifolia, Richardia grandiflora, Sebastiana hispida, Tocoyena formosa, Zanthoxylum rigidum, and Sesbania virgata (fruit extract) showed acaricidal activity against the larval stage of Rhipicephalus (Boophilus) microplus higher than 95% at a 40% (w/v) concentration, while Hippocratea volubilis and Randia armata showed moderate efficacy and Croton glandulosus and Senna obtusifolia had no effect. The M. latifolia, A. hirta, R. grandiflora, and A. latissima raw extracts were evaluated for their activity against adults, and only A. hirta showed an efficacy close to 90%. Eighteen extracts had an efficacy of up to 95% against larvae at a 40% concentration, seven extracts were effective at 20%, and only one (Sebastiana hispida) was effective at a 5% concentration.

Multispecies resistance of cattle gastrointestinal nematodes to long-acting avermectin formulations in Mato Grosso do Sul.

The use of long-acting avermectins (AVMs) in cattle to treat infections with gastrointestinal nematodes was common in Brazil until its prohibition by state authorities. The prohibition; however, was rescinded in 2015, but a scientific discussion of the pros and cons of the use of these formulations is necessary. We evaluated the levels of resistance to 1.0 and 3.5% doramectin and to 3.15% ivermectin in cattle. The worms in animals treated with 3.5% doramectin were characterized by the suppression of oviposition and by a higher proportion of adult females carrying no eggs. Haemonchus placei, Cooperia punctata, C. pectinata, C. spatulata, and Oesophagostomum radiatum were resistant to the above compositions. The administration of long-acting AVM formulations did not result in a higher efficacy against these helminth populations.

Eimeria species in young and adult sheep raised under intensive and / or semi-intensive systems of a herd from Umuarama city, Parana State, Brazil

The present study aimed to identify Eimeria species in young and adult sheep raised under intensive and / or semi-intensive systems of a herd from Umuarama city, Parana State, Brazil using the traditional diagnostic methods and to correlate the infection level/types of infection in the different age/system in this herd. Fecal samples were collected from the rectum of 210 sheep and were subjected to laboratory analysis to differentiate the species. Furthermore, animals were observed to determine the occurrences of the clinical or subclinical forms of eimeriosis. Out of the 210 collected fecal samples, 147 (70%) were positive for Eimeria oocysts, and 101 (47.86%) belonged to young animals that were raised under intensive and / or semi-intensive farming systems. Oocysts from 9 species of Eimeria parasites were identified in the sheep at the following prevalence rates: E. crandallis, 50.0%; E. parva, 21.6%; E. faurei, 8.1%; E. ahsata, 8.1%; E. intricata, 5.4%; E. granulosa , 2.7%; E. ovinoidalis , 2.0%; E. ovina , 1.3%; and E. bakuensis , 0.6%. There were no differences regarding the more frequent Eimeria species among the different ages of animals or between the different farming management systems. Based on these data, E. crandallis was the most prevalent, followed by E. parva and E. faurei species, regardless of the age. Higher parasitism was diagnosed in the young animals that were raised in a confinement regime, and the disease found in the herd was classified as subclinical. Further studies should be conducted in this herd, to verify if the eimeriosis subclinical can cause damage especially in young animals with a high level of infection.

Effect of creep-fed supplement on the susceptibility of pasture-grazed suckling lambs to gastrointestinal helminths

This study evaluated the effect of creep feeding a protein supplement on the susceptibility of suckling lambs to infection with gastrointestinal helminths. Male and female lambs were grazed on Brachiaria spp. pastures next to their mothers. Animals were allocated to one of two treatments: creep feeding (261g/d) and control (no supplementation). The trial period was the suckling of lambs during two years of study: May-October 2013 and March-July 2014. Supplementary creep feeding of lambs improved animal performance (P<0.05). Creep-fed lambs reached 18kg body weight in 64 d, but unsupplemented lambs required 77 d to reach the same weight. Lambs were susceptible to helminth infection during lactation; lambs in both treatments had high fecal egg counts (FECs), with means >1000 eggs per gram, as early as 45days of age, when the daily grazing time per animal increased. Creep feeding reduced the FECs of suckling lambs >60days of age in infections dominated by Haemonchus contortus. Totals of 20 and 48 anthelmintic treatments were administered to the supplemented and unsupplemented animals, respectively. The effect of this variable, however, was significant (P<0.05) only after 60days, when nine and 28 treatments had been administered to the supplemented and unsupplemented lambs, respectively. The number of strongyloid larvae recovered from the paddock did not differ significantly (P >0.05) between the two treatments, indicating similar challenges by infective larvae to both groups. The supplementation of lambs by creep feeding can thus be a strategy for the sustainable control of helminth infection, because it reduces the dependence on anthelmintic treatment.

First genetic identification of Cryptosporidium parvum subtype IIaA14G2R1in beef cattle in Brazil.

The presence of Cryptosporidium spp. in a cattle herd registered with an outbreak of diarrhea was investigated and the the molecular subtyping of Cryptosporidium parvum was characterized. Fecal samples from 85 Nellore beef cattle (Bos indicus) were collected and examined with Ziehl-Neelsen modified staining method. Fifty-four cattle (63.52%) had Cryptosporidium spp. oocysts in their feces. Fragments of genes encoding the 18S ribosomal RNA subunit and a 60-kDa glycoprotein (gp60) were amplified by nested PCR accomplished in the 11 most heavily parasitized samples, and the amplicons were sequenced. Eight of the 11 analyzed samples were positive for 18S rRNA sequences and identified monospecific infections with C. parvum. Seven samples were positive for gp60 and identified subtypes IIaA15G2R1 (6/11) and IIaA14G2R1 (1/11). This report is the first for C. parvum subtype IIaA14G2R1 in beef cattle in Brazil.

Retrospective study of epidemiological, clinical and pathological findings of bovine babesiosis in Mato Grosso do Sul, Brazil (1995 –2017)

Babesiosis is a tick-borne hemolytic disease caused by intraerythrocytic protozoan parasites of the genus Babesia. The present study describes epidemiological, clinical, and pathological features of cases of bovine babesiosis diagnosed over the last two decades in the state of Mato Grosso do Sul in central-western Brazil. Examination of a total of 5298 bovine samples resulted in a definitive or presumptive diagnosis of babesiosis in 41 cases from 33 outbreaks. Babesia bovis was the most common causal agent of bovine babesiosis, and animals of all ages were affected. Zebu cattle, which are more frequent in the study region than are taurine cattle, were the cattle most often affected. Although zebu cattle have been claimed to be highly tolerant, the present findings show that they can develop severe illness and die due to Babesia spp. infection. The disease was diagnosed almost every year over the time period covered in this study (1995-2017). However, the number of cases of bovine babesiosis diagnosed was low compared to the total number of cases reviewed. Veterinary practitioners that work on cattle farms should to be able to diagnose the disease in its early stages so that treatment can be initiated promptly and economic losses can be minimized.