This document summarizes a study on Hepatozoon canis infection in dogs in India. The study aimed to determine the molecular prevalence of H. canis, identify risk factors, and examine hematological changes in infected dogs. Blood samples from 225 dogs with anemia symptoms were collected and tested via microscopy, DNA extraction, PCR and analysis of hematological parameters. Results found H. canis gamonts in neutrophils and monocytes on blood smears and PCR identified the parasite DNA. Infected dogs showed significant decreases in hemoglobin and platelet counts compared to uninfected dogs, indicating anemia and thrombocytopenia. Risk factors like seasonal variation and ticks were also discussed.

1. MARIA CAMILA BEDOYA PIEDRAHITA
III SEMESTRE - UPB

2. SEMINARIO BIOLOGÍA
MOLECULAR
María Camila Bedoya Piedrahita| Esp. Lina Martínez| UPB

3. INTRODUCTION
• Hepatozoonosis caused by Hepatozoon canis is an important tick-borne
disease of dogs in tropical and subtropical regions throughout the world.
• Hepatozoon canis is a parasite that is considered to be one of the most
important vector-borne disease of canines with worldwide distribution.
• The principal vector of H. canis is the brown dog tick, Rhipicephalus
sanguineus.
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4. INTRODUCTION
• H. canis infection was found to be significantly associated with season
• The haematological alterations in the positive cases were lymphopenia,
anaemia, thrombocytopenia, relative neutrophilia, neutrophilic leucocytosis,
eosinophilia, monocytosis and lymphocytosis.
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5. GENERAL OBJECTIVE
• Recognize the molecular prevalence, risk factors and hematological changes
in a sample of dogs from Punjab, India thad had characteristic symptoms of
anemia and presented Hepatozoon Canis in the PCR test.
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6. MATERIALES Y MÉTODOS: MUESTRAS
Area geografica
•El estudio se realizó en el distrito de Ludhiana del estado de Punjab, India
•La aprobación y las pautas necesarias de Institute Animal Ethics Comité (AICE)
Muestras
•Se recolectó un total de 225 muestras de sangre en viales recubiertos con EDTA
•Los perros fueron seleccionados en la base de la historia y los signos clínicos sospechosos de estar infectados con protozoo
•Por un período de un año (2015-16)
Muestras
•1-2 ml se colocó en frascos con EDTA y el resto en anticoagulante libre
•Muestras se utilizaron para la preparación de sangre fina frotis y luego se mantuvieron a -20 ° C hasta la extracción de ADN
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7. MATERIALES Y MÉTODOS: MICROSCOPIA
Se realizó después de la tinción los
frotis de sangre finos preparados con
Giemsa
Examinado con aceite de inmersión
para para detectar los gamonts de H.
canis
La tasa de parasitemia se calculó con
el número de neutrófilos parasitados y
monocitos de 200 leucocitos en total.
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8. MATERIALES Y MÉTODOS: PRUEBAS
Extracción de ADN
3. Remover
proteínas
celulares
2.
Aislamiento
de ADN:
solución lisis
nuclear
1.
Purificación:
lisis celular y
nuclear
PCR
(Técnica de
amplificación
enzimática)
↑ Segmento
específico ADN
(gen 18S)
usando
polimerasa
Dos cebadores
que se unen al
ADN e inician
secuencia
(Hep-F y Hep-
R)
Producto de
cada ciclo es el
molde para
duplicar
DNTp
determina
secuencia de
AND
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Esta técnica sirve para amplificar un fragmento de ADN;
su utilidad es que tras la amplificación resulta mucho más
fácil identificar con una muy alta probabilidad, virus o
bacterias causantes de una enfermedad

9. MATERIALES Y MÉTODOS: ALTERACIONES
Muestrasdesangrese
usaronparadeterminar:
[Hb (g / dL)]
Total de globulos
rojos contadas TEC
Volumen de células
empaquetadas PCV
Recuento total de
leucocitos TLC
Recuento total de
plaquetas PLT
Recuento diferencial de se
realizó en 100x de un campo
brillante en Giemsa
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10. RESULTS
FIG 1. Giemsa stained thin blood
smear showing gamonts (arrows) of
H. canis in neutrophils of dog.

11. RESULTS
FIG 2. Field application of Hc-PCR
assay.
• Lane M: Generuler™ 100 bp
Ladder Plus
• Lane 1–5: Field collected
samples
• Lane N: Negative control
• Lane P: Positive control

12. TABLE 1. Comparison of various haematological parameters by ANOVA with Tukey’s multiple comparison test.
RESULTS
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13. RESULTS
TABLE 3. Distribution of
hepatozoonosis in dogs in
accordance with various risk factors.

14. DISCUSSION
AUTHOR DISCUSSION
[3] A. Gavazza, M. Bizzeti, R.
Papini
Regarding haematological alterations in the positive cases, Hb
levels were found to be significantly lower.
S
[30] L. Vojta, M. Mrljak, R. Beck
[33] H. Voyvoda, S. Pasa, A. Uner
Regarding the total platelet count, thrombocytopenia has been
reported by many workers in cases of hepatozoonosis in dogs
S
[13] S. Chhabra, S.K. Uppal, L.D.
Singla
[14] A.V. Mundim, I.A. Morais, M.
Tavares, M.C. Cury, M.J.
Anaemia,
neutrophilia, neutrophilic leucocytosis, lymphopenia,
thrombocytopenia, eosinophilia and monocytosis recorded as
the chief haematological alterations, in current study are also
consistent with the findings of other researchers
S
[19] N.K. Singh, Jyoti, M. Haque,
H. Singh, S.S. Rath
[21] H. Singh, N.K. Singh, S.S.
Rath
With reference to the age of the host, previous studies from the
same
region have reported non-significant variation regarding the
prevalence
of hepatozoonosis in dogs
S
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15. CONCLUSION
• In conclusion, the observed variations in the erythrogram and leukogram
parameters of the dogs in this study may also be related to parasitaemia, as
the patients with more decrease in hemoglobin have more. The decrease in
the level of proteins is due to the possible innsufficiency and kidney damage.
increase in leucites
• Environmental conditions provide favorable conditions for the survival and
spread of ticks (Rhipicephalus sanguineus). The most important factor of
propagation is summer.
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16. MAPA
CONCEPTUAL