Correlation Between Diarrhea Severity and Oocyst Count via Quantitative PCR or Fluorescence Microscopy in Experimental Cryptosporidiosis in Calves

Operario, Darwin J.; Bristol, Lauren S.; Liotta, Janice L.; Nydam, D.V.; Houpt, Eric R.

doi:10.4269/ajtmh.14-0488

Cited by 23 publications

(20 citation statements)

References 22 publications

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“…There was poor correlation between qPCR and microscopy for the 8 samples for which data from both methods were available, with qPCR detecting higher numbers of oocysts than microscopy with the exception of one sample (Eastern grey kangaroo 3). Increased sensitivity of qPCR and the estimation of much higher numbers of oocysts in faecal samples by qPCR versus microscopy has been previously reported [83]. A major limitation of qPCR is that the quantitative data generated are only as accurate as the standards used.…”

Section: Discussionmentioning

confidence: 99%

Zoonotic Cryptosporidium Species in Animals Inhabiting Sydney Water Catchments

et al. 2016

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Cryptosporidium is one of the most common zoonotic waterborne parasitic diseases worldwide and represents a major public health concern of water utilities in developed nations. As animals in catchments can shed human-infectious Cryptosporidium oocysts, determining the potential role of animals in dissemination of zoonotic Cryptosporidium to drinking water sources is crucial. In the present study, a total of 952 animal faecal samples from four dominant species (kangaroos, rabbits, cattle and sheep) inhabiting Sydney’s drinking water catchments were screened for the presence of Cryptosporidium using a quantitative PCR (qPCR) and positives sequenced at multiple loci. Cryptosporidium species were detected in 3.6% (21/576) of kangaroos, 7.0% (10/142) of cattle, 2.3% (3/128) of sheep and 13.2% (14/106) of rabbit samples screened. Sequence analysis of a region of the 18S rRNA locus identified C. macropodum and C. hominis in 4 and 17 isolates from kangaroos respectively, C. hominis and C. parvum in 6 and 4 isolates respectively each from cattle, C. ubiquitum in 3 isolates from sheep and C. cuniculus in 14 isolates from rabbits. All the Cryptosporidium species identified were zoonotic species with the exception of C. macropodum. Subtyping using the 5’ half of gp60 identified C. hominis IbA10G2 (n = 12) and IdA15G1 (n = 2) in kangaroo faecal samples; C. hominis IbA10G2 (n = 4) and C. parvum IIaA18G3R1 (n = 4) in cattle faecal samples, C. ubiquitum subtype XIIa (n = 1) in sheep and C. cuniculus VbA23 (n = 9) in rabbits. Additional analysis of a subset of samples using primers targeting conserved regions of the MIC1 gene and the 3’ end of gp60 suggests that the C. hominis detected in these animals represent substantial variants that failed to amplify as expected. The significance of this finding requires further investigation but might be reflective of the ability of this C. hominis variant to infect animals. The finding of zoonotic Cryptosporidium species in these animals may have important implications for the management of drinking water catchments to minimize risk to public health.

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Section: Discussionmentioning

confidence: 99%

Zoonotic Cryptosporidium Species in Animals Inhabiting Sydney Water Catchments

et al. 2016

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“…Infected calves typically excrete oocysts with faeces for about 2 weeks (Fayer et al, 1998;O'Handley et al, 1999). Under experimental conditions, the Cryptosporidium oocysts count in faeces rises a day before the onset of diarrhoea, peaks, and drops 2 days before the diarrhoea becomes less severe (Operario et al, 2015). In previous longitudinal studies, the highest number of Cryptosporidium oocysts were found during the second or third weeks of the calves' lives (Santín et al, 2008;Coklin et al, 2010).…”

Section: Introductionmentioning

confidence: 87%

Cryptosporidium outbreak in calves on a large dairy farm: Effect of treatment and the association with the inflammatory response and short-term weight gain

Niine

Dorbek-Kolin

Lassen

et al. 2018

Research in Veterinary Science

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Cryptosporidium spp. infections in neonatal dairy calves can cause diarrhoea and, in rare cases, death. The infection is usually self-limiting, but halofuginone lactate (HL) can be used prophylactically. Calves (n=144) in the study were born during a 2-month period on one farm. A total of 901 serum and 767 faecal samples were collected. Based on HL treatment, the calves were divided into 3 groups: I) not treated, II) treated incorrectly (treatment started >48h after birth, or lasted <7days), and III) treated correctly (started <48h after birth, and lasted ≥7days). Over the 3-month observation period, 14.6% (n=21) of the calves died, of which most (67%) had not been treated with HL. Correctly performed treatment of cryptosporidiosis significantly delayed the onset of oocysts shedding (P<0.001) and reduced haptoglobin (HP) and serum amyloid A (SAA) concentrations in the second week of life. HP concentration and HL treatment were negatively associated with weight gain at 3months of age. Cryptosporidium positive faecal samples were significantly (P<0.001) more likely to be diarrhoeic but Giardia or Eimeria positive samples were not. Correct prophylactic treatment with HL delayed the shedding of Cryptosporidium oocysts and improved survival, but was negatively associated with weight gain. Incorrect treatment had a low impact on mortality and resembled no treatment regarding the proportion of calves shedding oocysts. Acute phase response (APR) in the second week of life seemed to be positively associated with shedding high amounts of Cryptosporidium oocysts.

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“…qPCR for Cryptosporidium spp. 18S rRNA was performed using a previously described assay on the Applied Biosystems 7500-FAST plaCFCrm using an inhibitor-optimized master mix (Part 95134, Quantabio, Beverly, MA), 300 nM forward primer, 900 nM reverse primer and 120 nM of probe (labeled with FAM) [ 36 ].…”

Section: Methodsmentioning

confidence: 99%

Impact of confinement housing on study end-points in the calf model of cryptosporidiosis

et al. 2018

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BackgroundDiarrhea is the second leading cause of death in children < 5 years globally and the parasite genus Cryptosporidium is a leading cause of that diarrhea. The global disease burden attributable to cryptosporidiosis is substantial and the only approved chemotherapeutic, nitazoxanide, has poor efficacy in HIV positive children. Chemotherapeutic development is dependent on the calf model of cryptosporidiosis, which is the best approximation of human disease. However, the model is not consistently applied across research studies. Data collection commonly occurs using two different methods: Complete Fecal Collection (CFC), which requires use of confinement housing, and Interval Collection (IC), which permits use of box stalls. CFC mimics human challenge model methodology but it is unknown if confinement housing impacts study end-points and if data gathered via this method is suitable for generalization to human populations.MethodsUsing a modified crossover study design we compared CFC and IC and evaluated the impact of housing on study end-points. At birth, calves were randomly assigned to confinement (n = 14) or box stall housing (n = 9), or were challenged with 5 x 107 C. parvum oocysts, and followed for 10 days. Study end-points included fecal oocyst shedding, severity of diarrhea, degree of dehydration, and plasma cortisol.FindingsCalves in confinement had no significant differences in mean log oocysts enumerated per gram of fecal dry matter between CFC and IC samples (P = 0.6), nor were there diurnal variations in oocyst shedding (P = 0.1). Confinement housed calves shed significantly more oocysts (P = 0.05), had higher plasma cortisol (P = 0.001), and required more supportive care (P = 0.0009) than calves in box stalls.ConclusionHousing method confounds study end-points in the calf model of cryptosporidiosis. Due to increased stress data collected from calves in confinement housing may not accurately estimate the efficacy of chemotherapeutics targeting C. parvum.

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Correlation Between Diarrhea Severity and Oocyst Count via Quantitative PCR or Fluorescence Microscopy in Experimental Cryptosporidiosis in Calves

Cited by 23 publications

References 22 publications

Zoonotic Cryptosporidium Species in Animals Inhabiting Sydney Water Catchments

Zoonotic Cryptosporidium Species in Animals Inhabiting Sydney Water Catchments

Cryptosporidium outbreak in calves on a large dairy farm: Effect of treatment and the association with the inflammatory response and short-term weight gain

Impact of confinement housing on study end-points in the calf model of cryptosporidiosis

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