American Association of Bovine Practitioners Conference Proceedings
Anthelmintic resistance is a worldwide problem in all livestock systems, with most reports of res... more Anthelmintic resistance is a worldwide problem in all livestock systems, with most reports of resistance in cattle parasites being to the macrocyclic lactone (ML) drug class. Several prevalence studies performed internationally demonstrate that anthelmintic resistance in cattle parasites is a growing problem, however, no similar studies have been performed in the US. The primary objective of this study was to gain data on the prevalence of ML resistance in gastrointestinal nematode (GIN) parasites on beef cattle farms in Georgia. We also wanted to test the efficacy of combination anthelmintic therapy in these same herds, and use these data as a basis for educational outreach programs for beef producers in Georgia.
The microscopic examination of feces for the assess m nt of gastrointestinal parasite infection h... more The microscopic examination of feces for the assess m nt of gastrointestinal parasite infection has bee n a mainstay of clinical and research parasitology labs for many decades. Even with the widespread use of f cal egg counts (FECs) in the medical and scientific com munity, the routine use of FECs by farmers and prod ucers is quite limited in most areas of the world. The lack of use of this very valuable tool is most probably due to a lack of consistent and understandable information regard ing the simplicity and value of the FEC. Some of theinformation that one can obtain via the internet concerningFECs is of variable quality and clarity. Accurate information and training can help to correct the mi sunderstandings concerning the limitations of the F EC as well as expand its acceptance and use. The modified Mc Master FEC technique, which is one of the most wide ly used quantitative FEC methods in practice today, si mple to perform and when used as an adjunct to body condition score...
a r t i c l e i n f o Anthelmintic resistance of gastrointestinal nematode (GIN) parasites consti... more a r t i c l e i n f o Anthelmintic resistance of gastrointestinal nematode (GIN) parasites constitutes a major problem for livestock health and productivity around the world (Fleming et al., 2006; Kaplan and Vidyashankar, 2012). The common practice of intensive farming methods coupled with heavy reliance on anthelmintics has resulted in a serious escalation in the prevalence, distribution, GIN parasi ost severe increasing number of farms around the world experiencing tance to all classes of available anthelmintics (Howell 2008; Almeida et al., 2010; Sczesny-Moraes et al., 2010). M anthelmintic resistance in GIN parasites of cattle (Sutherland and Leathwick, 2011) and horses (von Samson-Himmelstjerna, 2012; Canever et al., 2013) is also being increasingly reported raising the level of concern in these hosts. In contrast to the situation with GIN, until recently the filarioid parasites, such as Dirofilaria immitis, were considered to be at a low risk for developing AR (Prichard, 2005)...
International Journal for Parasitology: Drugs and Drug Resistance, 2021
A population of Haemonchus contortus that was highly resistant to benzimidazoles and avermectin/m... more A population of Haemonchus contortus that was highly resistant to benzimidazoles and avermectin/milbemycins with a subpopulation that was resistant to levamisole, was replaced with a susceptible laboratory isolate of H. contortus in a flock of sheep. The anthelmintic susceptibility and population genetics of the newly established population were evaluated for 3.5 years using in vivo, in vitro, and molecular methods. Successful replacement of the resistant population with a susceptible population was confirmed using phenotypic and genotypic measurements; larval development assay indicated full anthelmintic susceptibility; albendazole treatment yielded 98.7% fecal egg count reduction; pyrosequence genotyping of single nucleotide polymorphisms in positions 167 and 200 of the isotype-1 beta tubulin gene were present at 0.0 and 1.7%, respectively; microsatellite genotyping indicated the background haplotype was similar to the susceptible isolate; and haplotypes of the isotype-1 beta tubu...
The levamisole-sensitive nicotinic acetylcholine receptor present at nematode neuromuscular junct... more The levamisole-sensitive nicotinic acetylcholine receptor present at nematode neuromuscular junctions is composed of multiple different subunits, with the exact composition varying between species. We tested the ability of two well-conserved nicotinic receptor subunits, UNC-38 and UNC-29, from Haemonchus contortus and Ascaris suum to rescue the levamisole-resistance and locomotion defects of Caenorhabditis elegans strains with null deletion mutations in the unc-38 and unc-29 genes. The parasite cDNAs were cloned downstream of the relevant C. elegans promoters and introduced into the mutant strains via biolistic transformation. The UNC-38 subunit of H. contortus was able to completely rescue both the locomotion defects and levamisole resistance of the null deletion mutant VC2937 (ok2896), but no C. elegans expressing the A. suum UNC-38 could be detected. The H. contortus UNC-29.1 subunit partially rescued the levamisole resistance of a C. elegans null mutation in unc-29 VC1944 (ok245...
International Journal for Parasitology: Drugs and Drug Resistance
Motility is a commonly used in vitro phenotype for assessing anthelmintic activity of candidate c... more Motility is a commonly used in vitro phenotype for assessing anthelmintic activity of candidate compounds, and for detecting anthelmintic resistance in nematodes. Third-stage larvae (L3) of parasitic nematodes are commonly used in motility-based assays because L3 are simple to obtain and can remain viable in storage for extended periods. To improve the measurement of motility of microscopic stages of nematodes, our laboratory developed the Worminator, which quantitatively measures motility of parasites. Using the Worminator, we compared the dose-response characteristics of several avermectin/milbemycin (AM) compounds using L3 from both AM-susceptible and AM-resistant Cooperia spp. (abamectin, doramectin, eprinomectin, ivermectin, moxidectin) and Haemonchus contortus (eprinomectin, ivermectin, moxidectin). Concentrations tested with the Worminator ranged from 0.156 to 40 μM. Differences in ECbetween AM-susceptible and AM-resistant isolates of Cooperia spp. and Haemonchus contortus were small, with resistance ratios ranging from 1.00 to 1.34 for Cooperia spp., 0.99 to 1.65 for Haemonchus contortus. Larval migration inhibition assays were conducted using the same isolates and were equally ineffective for detection of resistance with resistance ratios less than 2.0. These results contrast with those of the Larval Development Assay where we obtained a resistance ratio of 16.48 using the same isolates of Haemonchus contortus. Moreover, even at the highest concentration tested (40 μM), 100% inhibition of motility was never achieved and ECfor Worminator assays were more than 100× higher than peak plasma levels achieved in vivo following treatment. These data demonstrate that dose-response characteristics for inhibition of motility in L3 of gastrointestinal nematodes of livestock do not significantly differ for AM-susceptible and AM-resistant isolates. These data challenge the suitability of motility as a phenotype for detecting and measuring resistance to AM drugs in gastrointestinal nematodes of livestock.
Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc, 2016
Most small ruminant farms in tropical climates are plagued by Haemonchus contortus, a hematophago... more Most small ruminant farms in tropical climates are plagued by Haemonchus contortus, a hematophagous, abomasal parasite. Heavy burdens of this parasite can cause anemia, hypoproteinemia, weight loss, and mortality in susceptible animals. Haemonchus contortus is becoming a major health concern in New World camelids as well, namely llamas (Llama glama) and alpacas (Vicugna pacos), yet little research has been conducted regarding its prevalence or pathology in these species. Herein, we present a retrospective review of llamas and alpacas that were admitted to The University of Georgia Veterinary Teaching Hospital and Athens Diagnostic Laboratory between the years 2002 and 2013. Antemortem fecal egg count (FEC) estimates performed on 30 alpacas were negatively correlated with hematocrit, hemoglobin, and red blood cell count. Total protein was not significantly correlated with FEC. On postmortem examination, 55 of 198 camelids, including 2 from the aforementioned antemortem review, were i...
Levamisole is an anthelmintic drug that acts by activating nicotinic acetylcholine receptors at t... more Levamisole is an anthelmintic drug that acts by activating nicotinic acetylcholine receptors at the nematode neuromuscular junction and causing paralysis. We measured the in vitro effects of levamisole on the motility of Brugia malayi microfilariae; after 2 h incubation the apparent IC 50 was 2.68 mM. Lower drug concentrations, such as 1 mM, caused an immediate total paralysis that lasted for up to 1 h, but was completely reversed by 2 h of incubation. The 'recovered' parasites were still completely susceptible to application of a second nicotinic agonist, pyrantel.
International Journal for Parasitology: Drugs and Drug Resistance, 2013
For more than 20 years, anthelmintics of the macrocyclic lactone (ML) drug class have been widely... more For more than 20 years, anthelmintics of the macrocyclic lactone (ML) drug class have been widely and effectively used as preventives against the canine heartworm, Dirofilaria immitis. However, in recent years an increased number of lack of efficacy (LOE) cases are being reported, in which dogs develop mature heartworm infections despite receiving monthly prophylactic doses of ML drugs. While this situation is raising concerns that heartworms may be developing resistance to MLs, compelling evidence for this is still lacking. Resolution of this dilemma requires validated biological or molecular diagnostic assays, but, unfortunately, no such tests currently exist. To address this need, we developed and optimized a larval migration inhibition assay (LMIA) for use with D. immitis third-stage larvae. The LMIA was used to measure the in vitro dose-response of two ML drugs (ivermectin and eprinomectin) on a known MLsusceptible laboratory strain of D. immitis. A nonlinear regression model was fit to the dose-response data, from which IC 50 values were calculated; the mean IC 50 and 95% confidence interval for IVM was 4.56 lM (1.26-16.4 lM), greater than that for EPR at 2.02 lM (1.68-2.42 lM), and this difference was significant (p = 0.0428). The R 2 value for EPR assays (0.90) was also greater than that for IVM treatment (0.71). The consistency and reproducibility of the dose-response data obtained with this assay suggests that it may be a useful technique for investigating the relative susceptibilities to ML drugs in other D. immitis populations.
International journal for parasitology. Drugs and drug resistance, 2014
A major hindrance to evaluating nematode populations for anthelmintic resistance, as well as for ... more A major hindrance to evaluating nematode populations for anthelmintic resistance, as well as for screening existing drugs, new compounds, or bioactive plant extracts for anthelmintic properties, is the lack of an efficient, objective, and reproducible in vitro assay that is adaptable to multiple life stages and parasite genera. To address this need we have developed the "Worminator" system, which objectively and quantitatively measures the motility of microscopic stages of parasitic nematodes. The system is built around the computer application "WormAssay", developed at the Center for Discovery and Innovation in Parasitic Diseases at the University of California, San Francisco. WormAssay was designed to assess motility of macroscopic parasites for the purpose of high throughput screening of potential anthelmintic compounds, utilizing high definition video as an input to assess motion of adult stage (macroscopic) parasites (e.g. Brugia malayi). We adapted this ass...
An isolate of Haemonchus contortus, UGA/2004, highly resistant to benzimidazoles, levamisole, and... more An isolate of Haemonchus contortus, UGA/2004, highly resistant to benzimidazoles, levamisole, and ivermectin was isolated from sheep at the University of Georgia, and passaged through experimentally infected goats. We measured the expression of twenty-nine mRNAs encoding drug targets and P-glycoproteins (P-gps), comparing the results to a fully susceptible laboratory passaged isolate. Expression levels of some nicotinic acetylcholine receptor mRNAs were markedly different in UGA/2004. Levels of the Hco-acr-8b mRNA, encoding a truncated subunit, were very high in resistant L3, but undetectable in susceptible larvae, with expression of the full-length Hco-acr-8a mRNA also significant increased. Expression of Hco-unc-63 and Hco-unc-29.3 mRNAs was significantly reduced in the resistant larvae. Expression of the Hco-glc-3 and Hco-glc-5 mRNAs, encoding glutamate-gated chloride channel subunits, were slightly reduced in resistant larvae. We observed significant increases in the expression of the Hco-pgp-2 and Hcopgp-9 mRNAs in the UGA/2004 larvae, consistent with previous reports; we also saw a decrease in the levels of Hco-pgp-1 mRNA. Treatment of the larvae with ivermectin and moxidectin in vitro produced variable and inconsistent changes in P-gp mRNA levels. The sequences of the -tubulin isotype 1 mRNAs showed that the resistant larvae had a resistance-associated allele frequency of >95% at codon 200 and ∼40% and codon 167. No changes at codon 198 were present. The presence of the truncated acr-8b mRNA may be a reliable indicator of levamisole resistance, but complex changes in gene expression associated with macrocyclic lactone resistance make the identification of a single genetic marker for this resistance difficult.
Gastrointestinal nematodes of the family Trichostrongylidae are the most important parasites of s... more Gastrointestinal nematodes of the family Trichostrongylidae are the most important parasites of sheep, goats and other ruminants worldwide. Of this group, Haemonchus contortus is usually the most damaging species, particularly in warmer climates or during the summer. It is therefore useful to be able to rapidly differentiate infections with this nematode from other, less pathogenic, species. However, aside from Nematodirus spp., there are only subtle differences between the egg morphology within the trichostrongyles, making it very difficult to identify eggs to the species level. It has been shown previously that peanut agglutinin specifically binds to Haemonchus eggs and not those of other trichostrongyle species. By using this lectin conjugated to fluorescein isothiocyanate, binding to Haemonchus eggs can be visualized under ultraviolet illumination. We adapted this test using eggs purified by routine sugar centrifugation methods and evaluated 26 diagnostic samples from ruminants ...
American Association of Bovine Practitioners Conference Proceedings
Anthelmintic resistance is a worldwide problem in all livestock systems, with most reports of res... more Anthelmintic resistance is a worldwide problem in all livestock systems, with most reports of resistance in cattle parasites being to the macrocyclic lactone (ML) drug class. Several prevalence studies performed internationally demonstrate that anthelmintic resistance in cattle parasites is a growing problem, however, no similar studies have been performed in the US. The primary objective of this study was to gain data on the prevalence of ML resistance in gastrointestinal nematode (GIN) parasites on beef cattle farms in Georgia. We also wanted to test the efficacy of combination anthelmintic therapy in these same herds, and use these data as a basis for educational outreach programs for beef producers in Georgia.
The microscopic examination of feces for the assess m nt of gastrointestinal parasite infection h... more The microscopic examination of feces for the assess m nt of gastrointestinal parasite infection has bee n a mainstay of clinical and research parasitology labs for many decades. Even with the widespread use of f cal egg counts (FECs) in the medical and scientific com munity, the routine use of FECs by farmers and prod ucers is quite limited in most areas of the world. The lack of use of this very valuable tool is most probably due to a lack of consistent and understandable information regard ing the simplicity and value of the FEC. Some of theinformation that one can obtain via the internet concerningFECs is of variable quality and clarity. Accurate information and training can help to correct the mi sunderstandings concerning the limitations of the F EC as well as expand its acceptance and use. The modified Mc Master FEC technique, which is one of the most wide ly used quantitative FEC methods in practice today, si mple to perform and when used as an adjunct to body condition score...
a r t i c l e i n f o Anthelmintic resistance of gastrointestinal nematode (GIN) parasites consti... more a r t i c l e i n f o Anthelmintic resistance of gastrointestinal nematode (GIN) parasites constitutes a major problem for livestock health and productivity around the world (Fleming et al., 2006; Kaplan and Vidyashankar, 2012). The common practice of intensive farming methods coupled with heavy reliance on anthelmintics has resulted in a serious escalation in the prevalence, distribution, GIN parasi ost severe increasing number of farms around the world experiencing tance to all classes of available anthelmintics (Howell 2008; Almeida et al., 2010; Sczesny-Moraes et al., 2010). M anthelmintic resistance in GIN parasites of cattle (Sutherland and Leathwick, 2011) and horses (von Samson-Himmelstjerna, 2012; Canever et al., 2013) is also being increasingly reported raising the level of concern in these hosts. In contrast to the situation with GIN, until recently the filarioid parasites, such as Dirofilaria immitis, were considered to be at a low risk for developing AR (Prichard, 2005)...
International Journal for Parasitology: Drugs and Drug Resistance, 2021
A population of Haemonchus contortus that was highly resistant to benzimidazoles and avermectin/m... more A population of Haemonchus contortus that was highly resistant to benzimidazoles and avermectin/milbemycins with a subpopulation that was resistant to levamisole, was replaced with a susceptible laboratory isolate of H. contortus in a flock of sheep. The anthelmintic susceptibility and population genetics of the newly established population were evaluated for 3.5 years using in vivo, in vitro, and molecular methods. Successful replacement of the resistant population with a susceptible population was confirmed using phenotypic and genotypic measurements; larval development assay indicated full anthelmintic susceptibility; albendazole treatment yielded 98.7% fecal egg count reduction; pyrosequence genotyping of single nucleotide polymorphisms in positions 167 and 200 of the isotype-1 beta tubulin gene were present at 0.0 and 1.7%, respectively; microsatellite genotyping indicated the background haplotype was similar to the susceptible isolate; and haplotypes of the isotype-1 beta tubu...
The levamisole-sensitive nicotinic acetylcholine receptor present at nematode neuromuscular junct... more The levamisole-sensitive nicotinic acetylcholine receptor present at nematode neuromuscular junctions is composed of multiple different subunits, with the exact composition varying between species. We tested the ability of two well-conserved nicotinic receptor subunits, UNC-38 and UNC-29, from Haemonchus contortus and Ascaris suum to rescue the levamisole-resistance and locomotion defects of Caenorhabditis elegans strains with null deletion mutations in the unc-38 and unc-29 genes. The parasite cDNAs were cloned downstream of the relevant C. elegans promoters and introduced into the mutant strains via biolistic transformation. The UNC-38 subunit of H. contortus was able to completely rescue both the locomotion defects and levamisole resistance of the null deletion mutant VC2937 (ok2896), but no C. elegans expressing the A. suum UNC-38 could be detected. The H. contortus UNC-29.1 subunit partially rescued the levamisole resistance of a C. elegans null mutation in unc-29 VC1944 (ok245...
International Journal for Parasitology: Drugs and Drug Resistance
Motility is a commonly used in vitro phenotype for assessing anthelmintic activity of candidate c... more Motility is a commonly used in vitro phenotype for assessing anthelmintic activity of candidate compounds, and for detecting anthelmintic resistance in nematodes. Third-stage larvae (L3) of parasitic nematodes are commonly used in motility-based assays because L3 are simple to obtain and can remain viable in storage for extended periods. To improve the measurement of motility of microscopic stages of nematodes, our laboratory developed the Worminator, which quantitatively measures motility of parasites. Using the Worminator, we compared the dose-response characteristics of several avermectin/milbemycin (AM) compounds using L3 from both AM-susceptible and AM-resistant Cooperia spp. (abamectin, doramectin, eprinomectin, ivermectin, moxidectin) and Haemonchus contortus (eprinomectin, ivermectin, moxidectin). Concentrations tested with the Worminator ranged from 0.156 to 40 μM. Differences in ECbetween AM-susceptible and AM-resistant isolates of Cooperia spp. and Haemonchus contortus were small, with resistance ratios ranging from 1.00 to 1.34 for Cooperia spp., 0.99 to 1.65 for Haemonchus contortus. Larval migration inhibition assays were conducted using the same isolates and were equally ineffective for detection of resistance with resistance ratios less than 2.0. These results contrast with those of the Larval Development Assay where we obtained a resistance ratio of 16.48 using the same isolates of Haemonchus contortus. Moreover, even at the highest concentration tested (40 μM), 100% inhibition of motility was never achieved and ECfor Worminator assays were more than 100× higher than peak plasma levels achieved in vivo following treatment. These data demonstrate that dose-response characteristics for inhibition of motility in L3 of gastrointestinal nematodes of livestock do not significantly differ for AM-susceptible and AM-resistant isolates. These data challenge the suitability of motility as a phenotype for detecting and measuring resistance to AM drugs in gastrointestinal nematodes of livestock.
Journal of veterinary diagnostic investigation : official publication of the American Association of Veterinary Laboratory Diagnosticians, Inc, 2016
Most small ruminant farms in tropical climates are plagued by Haemonchus contortus, a hematophago... more Most small ruminant farms in tropical climates are plagued by Haemonchus contortus, a hematophagous, abomasal parasite. Heavy burdens of this parasite can cause anemia, hypoproteinemia, weight loss, and mortality in susceptible animals. Haemonchus contortus is becoming a major health concern in New World camelids as well, namely llamas (Llama glama) and alpacas (Vicugna pacos), yet little research has been conducted regarding its prevalence or pathology in these species. Herein, we present a retrospective review of llamas and alpacas that were admitted to The University of Georgia Veterinary Teaching Hospital and Athens Diagnostic Laboratory between the years 2002 and 2013. Antemortem fecal egg count (FEC) estimates performed on 30 alpacas were negatively correlated with hematocrit, hemoglobin, and red blood cell count. Total protein was not significantly correlated with FEC. On postmortem examination, 55 of 198 camelids, including 2 from the aforementioned antemortem review, were i...
Levamisole is an anthelmintic drug that acts by activating nicotinic acetylcholine receptors at t... more Levamisole is an anthelmintic drug that acts by activating nicotinic acetylcholine receptors at the nematode neuromuscular junction and causing paralysis. We measured the in vitro effects of levamisole on the motility of Brugia malayi microfilariae; after 2 h incubation the apparent IC 50 was 2.68 mM. Lower drug concentrations, such as 1 mM, caused an immediate total paralysis that lasted for up to 1 h, but was completely reversed by 2 h of incubation. The 'recovered' parasites were still completely susceptible to application of a second nicotinic agonist, pyrantel.
International Journal for Parasitology: Drugs and Drug Resistance, 2013
For more than 20 years, anthelmintics of the macrocyclic lactone (ML) drug class have been widely... more For more than 20 years, anthelmintics of the macrocyclic lactone (ML) drug class have been widely and effectively used as preventives against the canine heartworm, Dirofilaria immitis. However, in recent years an increased number of lack of efficacy (LOE) cases are being reported, in which dogs develop mature heartworm infections despite receiving monthly prophylactic doses of ML drugs. While this situation is raising concerns that heartworms may be developing resistance to MLs, compelling evidence for this is still lacking. Resolution of this dilemma requires validated biological or molecular diagnostic assays, but, unfortunately, no such tests currently exist. To address this need, we developed and optimized a larval migration inhibition assay (LMIA) for use with D. immitis third-stage larvae. The LMIA was used to measure the in vitro dose-response of two ML drugs (ivermectin and eprinomectin) on a known MLsusceptible laboratory strain of D. immitis. A nonlinear regression model was fit to the dose-response data, from which IC 50 values were calculated; the mean IC 50 and 95% confidence interval for IVM was 4.56 lM (1.26-16.4 lM), greater than that for EPR at 2.02 lM (1.68-2.42 lM), and this difference was significant (p = 0.0428). The R 2 value for EPR assays (0.90) was also greater than that for IVM treatment (0.71). The consistency and reproducibility of the dose-response data obtained with this assay suggests that it may be a useful technique for investigating the relative susceptibilities to ML drugs in other D. immitis populations.
International journal for parasitology. Drugs and drug resistance, 2014
A major hindrance to evaluating nematode populations for anthelmintic resistance, as well as for ... more A major hindrance to evaluating nematode populations for anthelmintic resistance, as well as for screening existing drugs, new compounds, or bioactive plant extracts for anthelmintic properties, is the lack of an efficient, objective, and reproducible in vitro assay that is adaptable to multiple life stages and parasite genera. To address this need we have developed the "Worminator" system, which objectively and quantitatively measures the motility of microscopic stages of parasitic nematodes. The system is built around the computer application "WormAssay", developed at the Center for Discovery and Innovation in Parasitic Diseases at the University of California, San Francisco. WormAssay was designed to assess motility of macroscopic parasites for the purpose of high throughput screening of potential anthelmintic compounds, utilizing high definition video as an input to assess motion of adult stage (macroscopic) parasites (e.g. Brugia malayi). We adapted this ass...
An isolate of Haemonchus contortus, UGA/2004, highly resistant to benzimidazoles, levamisole, and... more An isolate of Haemonchus contortus, UGA/2004, highly resistant to benzimidazoles, levamisole, and ivermectin was isolated from sheep at the University of Georgia, and passaged through experimentally infected goats. We measured the expression of twenty-nine mRNAs encoding drug targets and P-glycoproteins (P-gps), comparing the results to a fully susceptible laboratory passaged isolate. Expression levels of some nicotinic acetylcholine receptor mRNAs were markedly different in UGA/2004. Levels of the Hco-acr-8b mRNA, encoding a truncated subunit, were very high in resistant L3, but undetectable in susceptible larvae, with expression of the full-length Hco-acr-8a mRNA also significant increased. Expression of Hco-unc-63 and Hco-unc-29.3 mRNAs was significantly reduced in the resistant larvae. Expression of the Hco-glc-3 and Hco-glc-5 mRNAs, encoding glutamate-gated chloride channel subunits, were slightly reduced in resistant larvae. We observed significant increases in the expression of the Hco-pgp-2 and Hcopgp-9 mRNAs in the UGA/2004 larvae, consistent with previous reports; we also saw a decrease in the levels of Hco-pgp-1 mRNA. Treatment of the larvae with ivermectin and moxidectin in vitro produced variable and inconsistent changes in P-gp mRNA levels. The sequences of the -tubulin isotype 1 mRNAs showed that the resistant larvae had a resistance-associated allele frequency of >95% at codon 200 and ∼40% and codon 167. No changes at codon 198 were present. The presence of the truncated acr-8b mRNA may be a reliable indicator of levamisole resistance, but complex changes in gene expression associated with macrocyclic lactone resistance make the identification of a single genetic marker for this resistance difficult.
Gastrointestinal nematodes of the family Trichostrongylidae are the most important parasites of s... more Gastrointestinal nematodes of the family Trichostrongylidae are the most important parasites of sheep, goats and other ruminants worldwide. Of this group, Haemonchus contortus is usually the most damaging species, particularly in warmer climates or during the summer. It is therefore useful to be able to rapidly differentiate infections with this nematode from other, less pathogenic, species. However, aside from Nematodirus spp., there are only subtle differences between the egg morphology within the trichostrongyles, making it very difficult to identify eggs to the species level. It has been shown previously that peanut agglutinin specifically binds to Haemonchus eggs and not those of other trichostrongyle species. By using this lectin conjugated to fluorescein isothiocyanate, binding to Haemonchus eggs can be visualized under ultraviolet illumination. We adapted this test using eggs purified by routine sugar centrifugation methods and evaluated 26 diagnostic samples from ruminants ...
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