Simpozion Medical CYF Medical Diagnosticul Dirofilariozei cyfmedical@gmail.com , pentru diagnostic metode de diagnostic si teste sunati la 0723318308.

1. Dirofilaria infections in animals and
humans
Pathogenesis and clinical presentation: Dirofilaria
immitis versus Dirofilaria repens

Claudio Genchi, Med Vet, PhD, EVPC Dipl
Dept of Veterinary Science and Public Health, Università
degli Studi of Milano
Bucharest, June 14th, 2012

2. In spite of his denomination [heartworm
disease] is a respiratory [pulmonary] disease
In dogs, the heart is
usually involved in
the late stage of the
disease

3. Host response to HW infection
Cat: susceptible but
resisting host; usually not
microfilaraemic
unpredictable
Dog: usually microfilaraemic
reservoir
predictable disease

4. • Chronic disease
• Sometime acute
phases in the late
stage of the infection
• Clinical picture very
typical
• Acute symptoms
• Sometime chronic
disease
• No typical
symptoms

5. No lesions/symptoms for years
Inflammatory pulmonary
disease
Arterial pulmonary disease
(pulmonary hypertension,
“cor pulmonale”)
right heart congestive
failure
DOG Pathogenesis

6. Pulmonar disease
Vascular
disease
Parenchymal
disease
Cor polmonale

7. Vascular disease
• proliferative endarteritis
• thrombosis

8. 1995
Wolbachia pipiens is a Gram-negative bacterium
belonging to rickettesiales

9. Tetracycline in drinking water (0.3%) prevents infection by B.
pahangi and L. sigmodontis, but not by A. vitae, in gerbils
1971: Wolbachia and the antifilarial properties of
tetracycline. An untold story
Prompted by a serious outbreak of staphylococcal dermatitis affecting
the filarial infected and non-infected Mongolian jirds, in 1971 J.W.
McCall observed that tetracycline used in treating the condition had a
prophylactic affect against Brugia pahangi and Litomosoides
sigmodontis but not against Acanthocheilonema viteae. The results
were presented at the 48th Annual Meeting of the American Society of
Parasitologists (25-29 June 1973), but none of this data was published
until 10 years later.
The following studies showed that B. pahangi and L. sigmondis arer
Wolbachia infected while A. vieae is an uninfected species.

10. What we learned in the ‘90s
4. Wolbachia is present in
most filarial nematodes,
with a few exceptions in
non pathogenic species
such as Acantocheilonema
vitae
Wolbachia pipientis an “old/new” story

11. Onchocerca volvulus
Onchocerca ochengi
Onchocerca gutturosa
Onchocerca gibsoni
Thelazia lacrimalis
Dirofilaria repens
Dirofilaria immitis
Wuchereria bancrofti
Brugia pahangi
Brugia malayi
Litomosoides sigmodontis
Acanthocheilonema viteae
93
100
95
65
100
61
100
58
68
100
100
100
100
73
64
100
100
There is a consistency of filaria/Wolbachia phylogenies
Filaria phylogeny Wolbachia phylogeny
Modified from Casiraghi et al. 2001, Parasitology

12. Electron micrograph of an
embryo of D. immitis
containing two wolbachiae
Immunohistochemical staining
of W. pipientis in D. immitis
using Ab against WSP
Wolbachia is a widespread intracellular symbiont
of filarial nematodes and arthropods

13. Wolbachia pipientis: what we know today
Wolbachia is necessary for the host nematode
100% prevalence in infected species (lateral chords male
and females/reproductive tract females/larvae)
whole branches of filarial trees consist of infected species
consistency of host/Wolbachia phylogenies
vertical transmission

14. Wolbachia pipientis: what we know today
Tetracycline treatment of filarial-infected hosts inhibits:
embryogenesis
development of L3-adult
microfilarial production
long term survival of adults

15. Antibiotic treatments are effective both on Wolbachia and
on filarial nematodes development and embryogenesis,
causing prophylactic effects and infertility of adult females
(Genchi et al., 1998; Casiraghi et al. 2002)

16. Effects of antibiotic treatment on D. immitis
embryogenesis
From Sacchi et al. 2003, Parassitologia
control treated treated

17.  Inflammation
 Cytokine pattern and polarization of the immune
response
 Adverse reaction to chemotherapy
 Desensitization of innate immunity
 Defence against host response (e.g. through catalase
activity)
 the release of bacteria has been shown to be
associated with the up-regulation of pro-inflammatory
cytokines, neutrophil recruitment and an increase in
aspecific immunoglobulins
There is an overall consistency of data indicating that
Wolbachia plays an important role in the pathogenesis
of filarial disease throughout

18.  Adverse reactions to filaricidal therapy (ivermectin,
DEC) are associated with Wolbachia and/or its DNA
in the bloodstream and peak levels of Wolbachia
correlate with levels of pro-inflammatory cytokines.
Furthermore, a major surface protein of Wolbachia
(WSP) from D. immitis has been shown to provoke
chemiokinesis and IL-8 production in canine
neutrophils in vitro.
There is an overall consistency of data indicating that
Wolbachia plays an important role in the pathogenesis
of filarial disease throughout

19. The last two points (desensitization of innate
immunity and defence against host response)
open the question of the biological role of
Wolbachia in filarial nematodes:
Wolbachia “manipulates” the immune response,
thus allowing the long-term survival of the filariae
in immuno-competent hosts.

20. then …
Dirofilaria/Wolbachia symbiosis is friendly for
the parasite but is quite a foe for the host

21. Proliferative endarteritis: lesions are progressively
worsening during the infection and when worms are
dying
D. immitis in an asymptomatic dog:
the endothelium has no evident
damages
Proliferative endarteritis in a
dog death for
thromboembolism

22. Thrombosis
The thrombosis is a
consequence of the
platelet activation on the
sub-epithelial tissues
caused by pro- and
inflammatory products
released by the worms
(Wolbachia
endosymbionts) after
spontaneous or
pharmacological death

23. Spontaneous thromboembolism

24. Parenchymal disease
• Alveolar damage
• Eosinophilic pneumonia [very
severe in cats]

25. … as a consequence the heart disease
• increase of pulmonal
resistance
• cor polmonale
• right heart congestive
failure
LV
RV
To note, the right ventricle, as a
consequence of its anatomical
structure (pocket cling to left
ventricle), is adaptable to volume
overloading but not to pressure
overloading

26. Reduction of vascular stream (obstruction of
small arteries and capillaries)
+
loss of wall elasticity (severe arteritis)
Pulmonar hypertension
Increase of the post-charge

27. Post-charge overloading
Cor pulmonale (recruitment of collateral vessels)
right heart congestive failure

28. Microfilarial pathology
Mfs can embolise in
several organs, as the
kidney causing
glomerulonephritis
complicate by the
deposition of Ag/Ab
complexes.

29. Acute on chronic
Chronic respiratory disease
caval syndrome thromboembolism pneumothorax
Clinical Presentation

30. Clinical Presentation
no symptoms for years
unless exaggerated worm burden
and/or strenuous exercise
coughing
lipothymias
abdominal effusion

31. Apparently good health
[fat], but ascites
[abdominal effusion]
Clinical Presentation

32. right heart congestive failure
ascites
weight loss
disorexia
atrial fibrillation (> 15 kg bw)

33. Pneumothorax

34. Caval Syndrome
• Due to sudden rising of pulmonary
pressure (thromboembolism) and/or
cardiac output failure
• Heartworm displacement from pulmonary
arteries to the right cardiac chambers.

35. Caval Syndrome

36. Caval Syndrome
• Most frequently in small size dogs (< 11 kg)
3 main symptoms :
• Dyspnoea
• Cardiac murmur (thrill)
• Haemoglobinuria (haematuria)

37. Caval syndrome
Mechanical damage of RBC
for blood high pressure and
vortex

38. Caval syndrome

39. Cat
Where canine HW infection exists, feline HW infection exists
too!
HW prevalence in cats is 8-10% of prevalence in dogs.
In cats, HW infection is basically pulmonary. Clinically, the
infection can be asymptomatic, chronic or acute. The most
frequent signs are respiratory or digestive. Frequent
respiratory signs are dyspnoea, and tachypnoea and less
frequently, sneezing. Vomiting not associated with eating is
frequent but other symptoms such as diarrhoea are
occasionally described. Some affected cats also present
prostrated, anorectic, showing weight loss.

40. Dyspnoea Coughing
Vomiting
Hemoptysis Lipothymias
Coughing Vomiting
Weight loss
Diarrhoea

41. In those very rare cases where worms are located in the
right heart, a systolic cardiac murmur due to tricuspid valve
insufficiency and galloping cardiac rhythm are common
findings at auscultation.
Neurological signs such as ataxia, syncope, blindness, or
vestibular alterations, can be seen depending on the ectopic
localization of the migrating worms.
In acute cases, cats can die so quickly that owners are
usually not able to report any clinical signs before sudden
death.
Cat

42. Four year-old, female cat.
Sudden death, the owner told that
the cat was normal until the previous
evening and he found the cat dead
early in the morning without any
previous clinical sign.
At necropsy, one female adult worm
and signs of thrombembolism
Sudden death and hemoptysis

43. Clinical signs are usually more evident when young
parasites arrive in the pulmonary arteries (4 to 6 months
after infection) and later when adult worms die. Congestive
heart failure, common in dogs, is not frequently in cats.
Some infected cats recover spontaneously. However, even
in cases of chronic infection, there is always the risk of
reversion to the acute phase when adult parasites die. It is
associated with an intense pulmonary inflammatory reaction
in response to thromboembolism that is responsible for
pulmonary infarction and hemorrhage.
Circulatory collapse and respiratory failure usually follow.
Clinical signs at this stage can include dyspnea, cyanosis,
hypothermia, ataxia, hemoptysis and syncope.
Cat

44. 34
9
79%
21%
self cured
death
3
6
11
23
7%
14%
26%
53%
sudden death
died after 8-4 months
asymptomatic, self cured after 21-48 months
symptomatic, self cured 18-49 months
43 cats HW Ag and
ECHO positive

45. Cat
Feline HW disease is now recognized as a significant
pulmonary syndrome defined as Heartworm Associated
Respiratory Disease (HARD). The syndrome has been
observed in aborted infections too.
Clinical signs associated with HARD are anorexia,
lethargy, weight loss, coughing, rapid heart rate, vomiting,
diarrhoea, blindness, convulsions, collapse and sudden
death.

46. Cat: HARD
Both experimental and in filed studies have shown that the
HARD occurs even in cats with no adult parasites. Thus,
unlike in dogs, HW disease in cats should not be linked only
with adult worms and parenchymal disease but should be
associated with the arrival and the early death of immature
worms soon after they reach the pulmonary arteries.
HW-Ab positive cats, but free of adult HWs, have the same
type of pulmonary lesions, suggesting that even transient
infection with HW leaves cats with long-lasting pulmonary
pathology.

47. Prophilaxys Chemically abbreviated HW infection
infection
PulmonalyarteriolesAlveoli

48. When the worms reach the pulmonary arteries, acute
inflammatory responses at the vascular and parenchymal
level follow characterized by the activation of pulmonary
intravascular macrophages (PIM, abundant in cats but not in
dogs). The activation of PIMs is a unique and likely the most
important feature of HW infection in cats and is responsible
for the exacerbated pulmonary reaction.
At this stage, pulmonary radiographic findings include an
interstitial inflammatory pattern related to the increased
production of cytokines by macrophages.
Cat: HARD

49. Studying the pathogenesis of acute death syndrome in HW
infected cats, it has been shown that the most severe lung
reactions occurred in hypersensithyzed cats [IgE response]
exposed to dead worms or antigens from adult parasites.
The acute death syndrome is a consequence of an acute,
IgE mediated systemic anaphylactic reaction.
Cat

50. Although Dirofilaria immitis causes a very severe
disease in dogs and cats, there is increasing concern
about D. repens infection
 Owners require healthy pets
 D. repens can causes distress in infected animals
 In sensitized hosts the disease can be severe/ very
severe

51. Dirofilaria repens
 Most infections asymptomatic or accidentally diagnosed during
surgery
 Subcutaneous, cold, not painful and nodules movable on the
underlying tissues containing the adult parasite or mf
Courteously Z. Svobodova, Fac.
Med Vet, Brno, Czeck Republic
Dirofilaria repens infection: subcutaneous dirofilariosis

52. Worms are visulized by ECO as double linear parallel
hyperechoic structures similar but thinner than D. immitis
adult’s echoes
Dirofilaria repens worms can be visualized by
echography in cutaneous nodules and removed by a
mini-invasive procedure
Venco et al Int J Appl Vet Med 9:217-223 2011

53. D. repens scrotal nodule

54. Seldom, in severe infection, the clinical findings
[localized alopecia due to scratching and rubbing] and
generalized dermatitis seems to be mediated by
Wolbachia endosymbionts
Courtesy T Živičnjak et al, 2006