Investigation of Factors Associated with Subclinical Infections of Giardia duodenalis and Cryptosporidium canis in Kennel-Housed Dogs (Canis lupus familiaris).

Giardia duodenalis and Cryptosporidium spp. are zoonotic protozoal pathogens, spread by a fecal-oral route, which can infect a wide range of hosts including but not limited to dogs and humans. Giardia was recently estimated to be present in 37% to 50% of kennel-housed dogs. Cryptosporidium infections in kennel-housed dogs have been reported in 7% to 21% of the population. The goal of this study was to define demographic factors and fecal scores associated with positive screening test cases of Giardia and Cryptosporidium in kennel-housed laboratory dogs in the state of Texas. Fecal samples were collected from 153 clinically normal laboratory dogs at an academic research facility and a local laboratory dog supplier. We used 3 diagnostic tests evaluated in parallel to determine test positivity to each organism: a human point-of-care coproantigen test, a direct immunofluorescent assay, and an in-house polymerase chain reaction. Dogs were significantly more likely to test positive for Giardia (45%) than Cryptosporidium (7%) (P < 0.01). Dogs that were 18 mo of age or younger had 3 times the odds (P = 0.009) of subclinical Giardia infection compared with older dogs. We found no significant relationship between age and Cryptosporidium prevalence. Dogs with hard feces (fecal score 1-2) at the time of screening had 0.34 times lower odds ( P = 0.049) of testing positive for Giardia than dogs with normal feces, but no statistically significant relationship was found between fecal score and Cryptosporidium -positive test status. With these findings, we demonstrated the value of considering age and fecal score when choosing which dogs to screen for subclinical Giardia. Additional studies with larger sample sizes should be conducted to determine the relationship between age and fecal score and subclinical Cryptosporidium infection.

Comparison of 3 Diagnostic Tests for the Detection of Giardia and Cryptosporidium spp. in Asymptomatic Dogs (Canis lupis familiaris).

After detecting Giardia and Cryptosporidium infections and coinfections in 2 litters of puppies in our vivarium, our team realized that we needed a simple, quick, and economical point-of-care test for concurrent screening of asymptomatic dogs for both organisms. Periodic screening of colony dogs and of all dogs introduced into a colony can prevent the spread of Giardia and Cryptosporidium to immunologically naïve animals and help keep staff safe from these zoonotic organisms. To compare methods for diagnosing Giardia and Cryptosporidium spp. in dogs, we used a convenience sampling of feces from 2 popula- tions of dogs; samples were tested with a lateral-flow assay (QC), a commercially-available direct fluorescent assay (DFA), and an inhouse PCR test using established primers. QC results were analyzed in 2 ways: 1) relative to a reference standard that permitted comparative interpretation of DFA and PCR results; and 2) using Bayesian analysis for comparison independent of a reference standard. The QC test showed good specificity for the detection of Giardia according to both the reference standard (95%) and the Bayesian analysis (98%). Similarly, specificity of the QC for the detection of Cryptosporidium was 95% according to the reference standard and 97% according to Bayesian analysis. However, the sensitivity of the QC test was much lower for both Giardia (reference standard, 38%; Bayesian analysis, 48%) and Cryptosporidium (25% and 40%, respectively). This study demonstrates that the QC test can be used to detect both Giardia and Cryptosporidium in dogs and that positive results can be accepted with confidence, whereas negative tests should be confirmed through secondary testing methods.

Intrauterine infusion of latency-associated peptide (LAP) during early porcine pregnancy affects conceptus elongation and placental size.

In the pig, transforming growth factor beta (TGFB), TGFB receptors (TGFBRs), and integrins are present during the peri-implantation period. Latency-associated peptide (LAP), a part of latent TGFB, can bind to integrin heterodimers via its Arg-Gly-Asp (RGD) sequence; therefore, ligand-receptor interactions between TGFB and TGFBRs, along with LAP and integrin heterodimers, may be functional in mediating events supporting conceptus elongation and attachment. With the use of surgically implantable osmotic pumps, we were able to maintain pregnancy with the aim of mechanistically altering in vivo receptor-ligand interactions involving TGFB with TGFBRs and LAP with integrins during porcine pregnancy. Day 9 pregnant gilts received intrauterine infusions of LAP-RGD, a recombinant mutant of LAP (LAP-RGE), or vehicle control and were ovariohysterectomized on Day 13 or 24 of pregnancy. We hypothesized that intrauterine infusion of LAP-RGD would decrease downstream signaling of TGFB while increasing LAP-integrin interactions and that net effect would enhance conceptus survival and attachment early in the peri-implantation period but possibly increase the chance of abnormal placentation later in pregnancy. Additionally, we hypothesized that infusion of LAP-RGE would disrupt TGFB signals but not alter integrin signaling, and thus the net result would be decreased conceptus survival and abnormal development. Unexpectedly, LAP-RGD intrauterine infusions resulted in a reduction of conceptus elongation, whereas infusions of LAP-RGE permitted implantation and placentation but resulted in larger fetal weight, allantois length, and allantoic fluid volume. Results suggest TGFB and integrins are contributing factors in the regulation of conceptus elongation and placental and fetal size.

Transforming growth factor beta (TGFB) signaling is activated during porcine implantation: proposed role for latency-associated peptide interactions with integrins at the conceptus-maternal interface.

The process of implantation is mediated by a complex network of signaling and adhesive factors. In the pig, latent and active transforming growth factor beta (TGFB), TGFB receptors (TGFBR), and integrins (ITGs) are present during the peri-implantation period. TGFB signals via TGFBR and activates downstream effector SMAD proteins 2 and 3 (p-SMAD2/3). Latency-associated peptide (LAP), part of the latent TGFB complex, is known to bind to ITG heterodimers and activate TGFB. We hypothesize that active TGFBs and TGFBRs along with LAP and ITGs functionally interact at the conceptus-maternal interface to mediate events essential for conceptus development and attachment in pigs. Uteri and conceptuses from days 10, 12, 16, 20, and 24 pregnant gilts were immunostained for TGFB, LAP, and ITG subunits (ITGAV, ITGB1, ITGB3, ITGB5, ITGB6, and ITGB8). Activation of TGFBRs was evaluated by the presence of phosphorylated downstream effector SMAD2/3. Binding of LAP to ITGs was also evaluated using porcine trophectoderm cells. Abundant active TGFB was detected at the apical surfaces of epithelia at the conceptus-maternal interface, and p-SMAD2/3 was detected at both conceptus attachment and nonattachment sites during implantation. Separate aggregates of LAP, ITGB1, ITGB5, and later ITGB3 were detected at the porcine conceptus-maternal interface, and binding of LAP to ITGs on apical surfaces was demonstrated. Results suggest that functional LAP-ITG adhesion complexes support conceptus attachment and promote TGFB activation leading to TGFB interaction with TGFBR supporting events of porcine implantation.