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1. Dirofilaria infections in animals and
humans
New insights in the prevention and treatment of
canine and feline dirofilarial infections

Claudio Genchi, Med Vet, PhD, EVPC Dipl
Dept of Veterinary Science and Public Health, Università
degli Studi di Milano
Bucharest, June 14th, 2012

2. Searching for an effective
chemotherapy and prevention
Until the first half of 900’s, no effective treatment:
• Essential turpentine oil
• Intravenous injections of 0.40% formalin
• Antimony salts: till 11 injections
G. Zanotti: Of the degree of receptiveness, diagnosis and, antimony therapy
of canine filariosis. Profilassi 16: 79-87, 1943
The use of antimony salts was promptly given up for the
severe side effects

3. In 1947 Otto and Maren suggested the use of
thiacetarsamide (TCA) against heartworm infection.
They were the first who experimented the efficacy of
arsenical salts in HW infected dogs as a model for
treatment of Wuchereria brancoftii human infection. At
that time, Wuchereria infection was quite frequent in
American soldiers fighting in the Pacific area.
Otto GF, Maren TH. Possible use of an arsenical compound in threatment of
heartworm in dogs. Vet Med 1947, 42: 128
Searching for an
effective chemotherapy
and prevention

4. TCA [Caparsolate]: histolesivity, difficultly to manage
(2.2 mg/kg bw iv, 12 hours apart for 2 consecutive days),
toxicity and discontinuity in efficacy limited an effective
control of canine heartworm disease, mostly for severe
infections.
When melarsomine dihydrocloride was put on the
market (1997), the production of tiacetarsamide was
stopped.
Searching for an effective
chemotherapy and
prevention

5. TCA vs Melarsomine [Immiticide®]
TCA:
 high affinity for red cells [Dirofilaria worms are not
aematophagous]; the efficacy varies with the age and
the sex of worms.
 high content of arsenic
 histolesivity, hepato- and nephrotoxicity
Melarsomine dihydrocloride :
 higher affinity for blood plasma
 less content of arsenic, les hepato- and
nephrotoxicity
 Max blood concentration within about 150 min, 2-3
higher than TCA

6. The idea that it would be possible to prevent HW
larval development to adult parasite preventing canine
HW disease was developed at the beginning of
1960’s.
Kume et al. (1962) confirmed the concept of HW
prevention through daily administration of
diethylcarbamazine (DEC).
In the 90’s, macrocyclic lactones prevention was
extended to both feline HW infection and to
subcutaneous filarial infections (Dirofilaria repens).
Searching for an effective chemotherapy
and prevention

7. … before the discovery of macrocyclic lactones
and of its efficacy against Dirofilaria larval stages:
2 options
1. Diethylcarbamazine (DEC) citrate: 1.25 mg DEC
base/4.5 kg bw/day.
To be effective, the treatment must begin shortly before
the start of the transmission season and continue daily
for 2 months after the end of the transmission season.
The omission of only 2 or 3 consecutive doses can
compromise efficacy and cause a lapse of protection.
Side effects: vomiting, diarrhea, male infertility, severe
anaphylactic reaction in microfilaremic dogs.

8. In 1980, John W. McCall presented the first results on
the preventive efficacy of the ivermectin and concluded
… in terms of efficacy, safety and infrequency of
dosing, ivermectin looks promising as a
chemoprofilactic agent against canine heartworm.

9. 25 years after Kume’s paper (1962), the introduction on
the market of ivermectin (1987), milbemycin oxime (1990)
and, more recently, of selamectin and moxidectin, made
prevention possible through monthly administration of a
preventive drug.
From 2002, an injectable moxidectin formulation is on the
veterinary market in some European countries. The drug is
able to protect dogs against HW infection throughout the
entire transmission season following a single injection.
Searching for effective
chemotherapy and prevention

10. Nowadays, several macrocyclic lactones products such
as ivermectin, milbemycin oxime, selamectin and
moxidectin are available in Europe for the prevention
and control of canine and feline Dirofilaria infections.
IVM and MBO formulations are; SLM formulation is for
topical treatment (spot-on); moxidectin formulations are
oral, topical (spot on) and injectable.
All are safe [including ivermectin-sensitive dogs], a full
efficacy is requested by regulatory authorities worldwide
and they have retroactive activity [ML safe net or reach-
back effect].

11. Do not forget: we say prevention but it is
a larvicidal treatment: MLs monthly or
injectable
Treatment is effective against L3 and L4 larvae
Treatment should begin within 1 month from the
beginning of transmission by infected
mosquitoes and the last dose should be given 1
month after transmission ceases

12. Dose of preventative drugs for
canine HW prophylaxis
Macrocyclic lactone Min-max dose
Ivermectin
Milbemycin oxime
Moxidectin (tablets)¹
Moxidectin/imidacloprid spot on
Moxidectin injectable³
Selamectin (spot-on)²
6 - 12 µg/kg bw
0.5 - 0.99 mg/kg bw
3 - 6 µg/kg bw
2.5 – 6.25 mg/kg bw
0.17 mg/kg bw
6 - 12 mg/kg bw
¹, ² the efficacy lasts 60 days; ³ full risk season efficacy

14. Dose of preventive drugs for the
feline HW prophylaxis
Macrocyclic lactone Minimum dose
Ivermectin
Milbemycin oxime
Moxidectin/imidacloprid spot on
Selamectin (pour-on)¹
24 - 71.7 µg/kg bw
2 - 4 mg/kg bw
1 - 2 mg/kg bw
6 - 12 mg/kg bw
1 the efficacy lasts 60 days

15. Efficacy
Preventative Dog Cat
IVM
MBO
MOXI tablets
MOXI pour on
MOXI inject
SLM
D. immitis
D. immitis, Toxocara, Toxascaris,
Ancylostoma, Trichuris
D. immitis
D. immitis, D. repens, Toxocara,
Toxascaris, Ancylostoma,
Trichuris, Sarcoptes, Otodectes,
Demodex (control)
D. immitis, D. repens
D. immitis, D. repens, Toxocara,
Toxascaris, Ancylostoma,
Cyenocephalides, Sarcoptes,
Otodectes
D. immitis, Toxocara, Toxascaris,
Ancylostoma
D. immitis, Toxocara, Toxascaris,
Ancylostoma,
D. immitis, D. repens, Toxocara,
Toxascaris, Ancylostoma,
Otodectes
D. immitis, D. repens, Toxocara,
Toxascaris, Ancylostoma,
Cyenocephalides, Otodectes

16. Parasites
age (mths)
No. of
treatments
Efficacy
%
Vitality/motility of
recovered worms
IVM 6µg/kg/mth
MBO 0.5 mg /kg/mth
SLM 6 mg/top/mth
3
4 - 5
7
8
3 - 4
3
adulti
13
12-31
29
16
12-14
12
18
98
95-99
95
56
41-97
98.5
39
abnormal
abnormal
abnormal
abnormal
normal
abnormal
abnormal
from Guerrero, McCall e Genchi, 2002
Reach-back efficacy of macrocyclic lactones

17. HW adulticide treatment: melarsomine
dihydrocloride
• Two intramuscular injections of 2.5
mg / kg 24 hours apart (standard
regimen, not advised)
► Strongly advised
two step treatment by
giving one injection and
then administering the
standard pair of injections
at least 60 days later
One administration of melarsomine at the dose of 2.5
mg/kg kills about 90% of male worms and 10% of female
worms resulting therefore in 50 % reduction of the worm
burden (which is safer in terms of embolism and shock).
A 10% death rate can be
expected in dogs with High
risk of thromboembolic
complications when 2
doses are given initially.

18. Given strictly intramuscularly, deep in the lumbar
(back) muscle. Do not administer at any other site. No
more than 2 ml in the same site. Alternate sides. Use
thin needles (21-22 gauge). Change the needle for the
injection (to avoid subcutaneous tissues contamination)

19. Pulmonary thromboembolism is an inevitable
consequence of a successful adulticide therapy. If
several worms die widespread pulmonary thrombosis
frequently develops. Mild thromboembolism may be
clinically unapparent, but in severe cases life
threatening respiratory distress can occur.
Watch for:
Coughing, Dyspnoea
Fever
Hemoptysis, Lipothymias
The most critical time is 7-10 days following a melarsomine
treatment but pulmonary thromboembolism can occur
anytime in the following month

20. ANCILLARY THERAPY
• REST
• Calcium heparine
• Prednisolone
• Aspirine

21. REST• After treatment, the patient
must be strictly confined for
one month following the
final treatment.
• No walks, no running
around.
• The dog must live the
indoors (or in cage in
selected cases). The
reason for this is that it is
the most important thing to
minimize embolism-related
problems.

22. Calcium heparin
 50-100 UI s.c. 3 times daily starting 1-2
weeks prior to and continuing for 4-6 weeks
after adulticidal treatment
NO

23. Prednisolone
 0.5 mg/kg every other day from 5 th to
10 th day post adulticide treatment

24. Even though not recommended, IVM administered orally
at the prophylactic dose of 6 µg/kg monthly throughout the
year for a period of at least 2-2.5 years has been shown to
kill adult parasites.
To note that throughout this period the infection would
persist and pathology continues to worsen. Furthermore
the long-term use of a macrocyclic lactone in heartworm
positive dogs is the potential for selection of resistant sub-
populations of heartworms.
This regimen should be restricted to selected cases,
excluding active dogs, working dogs and heavily infected
dogs. X-ray examination should be performed every 4-5
months throughout the treatment period to monitor the
pulmonary patterns.
What about the use of MLs as an adulticide
drug?

29. Recently, it has been shown that a combination of
ivermectin at 6 µg/kg given every 15 days for 180 days
and doxycycline at 10 mg/kg once daily for 30 days is
well tolerated, has good adulticide efficacy and reduces
the risk of thromboembolism. Exercise should be rigidly
restricted for the duration of the treatment process. An
antigen test should be performed every 6 months and the
combination treatment continued until two consecutive
negative heartworm antigen tests have been obtained.
Anecdotal reports on other macrocyclic lactones with
adulticidal properties suggest similar results but no
confirmatory studies have been published.
IVM plus Doxy

30. Surgical heartworm removal via jugular
vein in the dog
Why, when and wherefore…

31. Surgical intervention is advised when several worms
have been displaced into the right cardiac chambers
producing sudden onset of caval syndrome.
It can be accomplished under general anesthesia with
flexible alligator forceps introduced via the jugular vein
aided by fluoroscopic guidance which gives access
not only to the right cardiac chambers but also to the
major pulmonary arteries.

32. Whenever …
 Caval Syndrome
(compulsory!)
 Dogs harboring large
worm burden at
ECHO examination
(advised)

33. Flexible Alligator Forceps® Fujinon

34. SURGICAL TOOLS
 flexible Alligator Forceps
 fluoroscopy
 basic suturing tools
 2 haemostatic clips

35. Anaestetic protocol
 Ketamin hydrocloride 5 mg/kg i.v.
 Diazepam 0,25 mg/kg i.v.
 Oxigen (mask)

36. technique
main pulmonary artery

37. Surgical technique

38. Surgical technique

39. Surgical technique

40. Surgical technique

41. EFFICACY
negativization
of Ag test four months after surgery
 Caval Syndrome 78%
 Chronic Cases 65 %
Average of removed heartworms 90%

42. In cats, adultice therapy with melarsomine is not advised.
If a cat does not display overt clinical signs despite
radiographic evidence of pulmonary vascular/interstitial
lung disease consistent with HARD, it may be advised to
allow time for self-cure. Subclinical cases can be
monitored periodically (6- to 12-month intervals) by
antibody and antigen re-testing and thoracic xR.
In those cats destined to recover, regression of xR signs
and especially seroconversion of a positive antigen test to
negative status provide evidence that the period of risk
probably has passed.
Adulticide treatment of feline HW infection

43. Prednisone in diminishing doses is often an effective
medical support for infected cats with radiographic
evidence of lung disease, whether or not they are
symptomatic. Also, this should be initiated whenever
antibody-and/or antigen-positive cats display clinical signs.
An empirical oral regimen is 2 mg/kg body weight/day,
declining gradually to 0.5 mg/kg every other day by 2
weeks and then discontinued after an additional 2 weeks.
At that time, the effects of treatment should be
reassessed based on the clinical response and/or thoracic
radiography.
This treatment may be repeated in cats with recurrent
clinical signs
Adulticide treatment of feline HW infection

44. Compound Formulation Dose Safety Efficacy
Melarsomine
dihydrochloridre
Injectable
deep in the
muscles
2.5 mg
24 hrs
apart
Toxicity,
possible side
effects if the
compounds is
injected in
connective
tissues
NO
efficacy
D. repens adulticide
treatment, is it possible?

45. Adult worms can be removed using a 19 Gauge
needle, connected to a vacuum syringe

46. Compound Formulation Dose Safety
Ivermectin
Selamectin
Moxydectin
plus
Doxycycline
tablets/
chawable
topical
topical
Per os
6 mcg/kg 12-24 mths
60 mg/kg 12-24 mths
1 mg/kg 6-12 mths
10 mg/kg daily
6-8 weeks/intervals
2-3 weeks
Good
Possible idiosyncratic
intolerance in single
subjects
Because D. repens severe clinical
signs can be as a consequence of
Mfs sensitization

47. Compound Formulation Dose Reference
Selamectin
Ivermectin
Moxidectin
Topical
Tablet per os
Chewable
Injectable SR
6 mg/kg bw
6 mcg/kg bw
0.17 mcg/kg bw
Genchi et al., 2002
Fok et al., 2009
Marconcini et al., 1986
Pollono et al., 1998
Rossi et al., 2004
Dirofilaria repens prevention in dogs
MLs [larvicidal treatment]: field data from owned dogs
observed comparing the treatment season with previous
data of prevalence

48. Formulation Dose Efficacy Reference
IVM
MOX
Tablet per os
Injectable SR
Topical
6 mcg/kg bw
12 mcg/kg bw
0.17 mcg/kg bw
2.5 mg/kg bw
87%
93%
100%
100%
Cancrini et al,1989
Genchi et al, 2010
Genchi et al, 2012
Dirofilaria repens prevention in dogs
MLs [larvicidal treatment]: experimental data

49. MOXI is more lipophilic than IVM and it is stored in the
fat, which may act as a drug reservoir. Compared to IVM,
MOXI has a higher distribution volume and a longer half-
life elimination. This may facilitate its distribution from the
blood stream to different tissues and longer residence
time for the drug in the body.
D. repens larval stages and the adult worms are
permanent residents in subcutaneous tissues, which are
rich of connective and fat tissues and the high lipophilic
nature of MOXI are probably the reason of the full
efficacy.
D. repens prevention: MOXI vs IVM

50. Canine and feline HW patent infection can be safely and
effectively prevent by the use of macrocyclic lactones, bot
throughout a monthly administration in the risk season or
throughout moxidectin sustained release formulation
injection.
From WAAVP, FDA and EMEA guidelines of veterinary
medicinal the efficacy of such a treatments must be
100%
Background

51. What is new about HW?
Canine HW infection: 2005
Most failures have been reported in HW-endemic states ... it is
unclear whether these are representative of the rare occurrences
of failure that have been in existence for a long time, but not
reported regularly, or whether there is a true increase
ineffectiveness … Results suggest that more comprehensive
reporting will provide FDA/CVM more accurate surveillance
information regarding efficacy. It will permit to better interpret both
incidence and severity of ineffectiveness and possible emerging
resistance and to convey this in any necessary updated labelling.
It also indicates that practitioners should return to a more
conservative testing schedule.

52. 2011
A controlled lab study was conducted to evaluate the efficacy of 4
commercial products. On Day −30, dogs were infected with 100 D.
immitis L3. On Day 0, each dog group was treated with
ivermectin/pyrantel (IVM 6.6 mcg/kg) or milbemycin oxime (500 mcg/Kg)
orally, or selamectin (60 mg/kg) solution or moxidectin/imidacloprid
topically (MOX 100mcg/kg). Group 5 dogs remained nontreated.
At necropsy (Day 120), all 8 dogs in the no treated group were infected
with adult D. immitis (34–70 worms/dog). One or more adult D. immitis
and/or worm fragments were recovered from dogs in each treatment
group (efficacy 95.6, 95.4 and 95.5% efficacy). No worms were
recovered from any of the 8 dogs in Group 4 resulting in 100% efficacy.

53. Results: Thirteen of 14 control dogs had adult HW detected at necropsy
with a geometric mean worm count of 22.3. One HW was found in 1 dog
in each of the MBO and IVM treatment groups.
Conclusions and Clinical Importance: Two currently approved
macrocyclic lactone HW preventives used at their labelled dose rates
were < 100% effective against a recent HW field isolate, supporting the
hypothesis that the effectiveness of a single dose of these preventives
can vary. This is important in guiding clients on expectations of product
effectiveness.
Experimental studies with a prouved lab resistant strain

54. The combination product of spinosad (anti-flea) and MBO
provides effective control of canine heartworms. A single
treatment at 30 days post infection showed high but incomplete
effectiveness against a HWs isolate that had been shown to be
partially refractory to treatment with marketed monthly
heartworm preventives. Three consecutive monthly treatments
provided complete control, providing support to the
recommendation that heartworm prophylaxis should be
maintained year round for optimal effectiveness.
Experimental studies with a prouved lab resistant strain

55. Resistant field strain

56. Knott mf always
positive
IVM 50-200 µg
MB 0.75-2 mg x
8 gg

57. D. immitis has exhibited a lower than expected heterozygosis and
worm populations were found to be relatively homogeneous. For
instance, the Japanese field samples were found to share several
common combined genotypes with US field samples. The
situation differs when laboratory isolates are compared with field
isolates from the same country or with field strains from other
countries (i.e. from US lab and US field and US lab and
Japanese field).
To note that the genetic polymorphism of D. immitis populations is
a consequence of many factors both of the mammalian host and
the invertebrate intermediate hosts (mosquito); however, all
previous data were obtained from dogs in areas that have been
intensively exposed to macrocyclic lactone preventatives for over
20 years (US, Japan and Italy). Nevertheless, the different field
isolates studied by did show identical gene sequences.
Towards resistance in Europe?

58. HW: last recommendations
Before the seasonal HW preventive treatment:
- Knott microfilariae, Ag test kit
Before and after adulticide HW treatment:
- Knott microfilariae, Ag test kit to assess the efficacy
The presence of resistant microfilariae must be carefully
checked to avoid the infection of mosquitoes and the
spreading of resistance

59.  Although Dirofilaria does not appear entirely
dependent on its bacterial symbiont Wolbachia, which
can be killed by a prolonged antibiotic treatment, the
clearing of bacteria from circulating microfilarie seems
to prevent infective larvae which develop in mosquitoes
from continuing their development in dogs.
 The combination of heartworm preventatives with
products designed to prevent mosquito blood-feeding
activity (repellents) during the heartworm transmission
season could be useful in protecting dogs from
infection and from ectoparasite infestations that often
occur in the same season.
HW: last recommendations

60. Dirofilaria repens: last recommendations
Before seasonal D. repens preventive treatment:
- Knott microfilariae
[No Ag test kit for filarial subcutaneous infections]
Microfilaraemic dogs must be treated against mfs with
macrocyclic lactones at the same dose to prevent patent
D. immitis infection for 3-12 moths [Knott examination until
negative results] to control the spreading of the infection
and preserve human health.
D. repens human infection is an emergent zoonosis in
Europe and human cases are increasing.

61. Mf clerance from blood stream
Ivermectin 6 mcg/kg
Milbemicine ossime 0.5
mg/kg
Selamectin 6 mg/kg
Moxidectin 2.5 mg/kg
Low efficacy (4-6 mth adm), mild reaction as a
consequence of the death of some mfs
Very effective, hypersensibility reactions in dogs
with high microfilaraemia
Low effecacy, no side effect after treatment in
microfilaraemic dogs
Efficacy after 4-6 moth administrations, no side
effects
Microfilaria treatment [mf clearance]
Before the prophylactic treatment with MBO products, dogs must be Knott
examined for circulating Mfs
Interceptor, Sentinel, [MBO + lufenuron], Trifexis [MBO + spinosad]

62. Starting and ending HW temperature
suitability in some European localities
2008
6-7 mth
treatment
May-Oct/Nov