**3. Breeds and tolerance to TBD in Apulia**

454 Health Management – Different Approaches and Solutions

2 days 9.60.9 13.01.0 42.04.1 43.73.0 13.50.9 31.11.34 4.51.0 7 days 8.71.1 12.60.9 39.71.9 46.26.0 14.72.2 31.71.17 5.41.6 15 days 9.00.9 12.50.7 39.52.0 44.12.8 14.00.5 31.71.5 5.62.8 21 days 9.40.8 11.60.7 35.71.5 38.23.32 12.41.0 32.52.1 5.31.7 30 days 9.21.2 11.20.5 35.41.9 38.74.9 12.41.5 31.71.4 6.12.5 45 days 9.91.2 10.80.8 35.52.0 36.13.8 11.01.4 30.51.9 7.52.1 2 months 9.80.8 10.70.4 34.92.1 35.63.0 10.90.9 30.61.3 7.81.7 3 months 9.30.8 10.50.4 32.71.4 35.43.1 11.41.0 32.20.9 6.81.8 4 months 9.11.2 10.30.6 32.61.9 36.23.1 11.41.0 31.50.8 8.12.0 7 months 7.80.5 9.10.5 29.91.4 38.41.3 11.70.6 30.60.8 7.91.9 9 months 7.90.7 9.40.8 30.11.3 38.12.6 11.90.9 31.21.5 7.31.4 12 months 7.50.3 9.40.5 28.51.5 38.02.1 12.40.6 32.81.3 8.91.4 15 months 7.60.4 9.30.4 28.41.5 37.52.5 12.20.7 32.71.3 8.81.4 18 months 7.80.3 9.30.3 28.60.7 36.81.0 12.00.5 32.60.7 8.61.4

Table 2. Least-square means (LSM) and standard errors (SE) of haematological values recorded for 22 Altamurana lambs controlled from birth to 18 months of age. Modified from

Pieragostini et al. (2000). RBC, Red Blood Cells; Hb, Haemoglobin; PCV, Packed Cell Volume; MCV, Mean Corpuscular Volume; MCH, Mean Corpuscular Haemoglobin,

Considering that reference data are mainly from breeds originally selected in northern European countries, when a comparison was made between 12 month-old Altamurana lambs and their northern counterparts, the erythrocytes of the Altamurana were fewer (7.5 *versus* 11.8 millions/l) but bigger (38.0 ft *versus* 26.5 ft) and full of haemoglobin (12.4 pg *versus* 9.3 pg). This is the same phenomenon encountered in Mediterranean/North-African

The overall pattern is suggestive of erythrocyte physiological effectiveness, which was confirmed by the perfect physical development of the subjects examined in this study. In the blood smears obtained at seven months of age, namely in full spring when lambs start to graze pastures, endoerythrocytic enzootic parasites (*Theileria* spp. and *Anaplasma* spp.) were recorded and then became a constantly occurring phenomenon as will be documented in the

MCHC, Mean Corpuscular Haemoglobin Content; WBC, White Blood Cells.

ovine blood picture as well as in the native Apulian adults.

following sections.

PCV (g/dl)

**Age** RBC

(106/l)

Hb (g/dl) **Haematological parameters** 

LSM±SE LSM±SE LSM±SE LSM±SE LSM±SE LSM±SE LSM±SE

MCH (pg)

MCHC (g/dl)

WBC (103/l)

MCV (fl)

Tick-borne diseases are of global importance to human and animal health and welfare. They are also responsible each year for dramatic economic losses which comprise direct losses from death of animals, loss of productivity and indirect losses due to the costs of control measures. In 1979, the amount of losses were estimated to be globally USD 7 billion (McCosker, 1979), but several reports on the economic costs of specific tick-borne diseases indicated that the earlier report is an underestimate (Jongejan & Uilenberg, 2004). There is a wide portfolio of measures which could be used to control tick-borne diseases among which both husbandry practices and host-related factors such as age, innate tolerance and breed are of great importance. Breeds whose historical breeding site is situated under the latitude of 41° show the ability to thrive in areas where tick borne diseases (TBD) are common. This trait, which can be defined as tolerance to TBD, is associated with the ability to resist the development of anemia in the face of infection.

A review on host resistance to tick borne diseases is documented in cattle (Correia de Almeida Regitano & Prayaga, 2010). As for other species, the case of the tolerance to tickborne diseases shown by the sheep and horse native to Apulia is emblematic (Pieragostini & Petazzi, 1999; Rubino et al., 2006). In southern Italy, and particularly in Apulia, pyroplasmosis represents a longstanding and heavy burden for every type of livestock farm (Ceci & Carelli, 1999). Previous work performed on Gentile di Puglia sheep found that blood smears for parasite detection revealed an overall positivity rate of 93% for tick borne parasites (TBP) (Pieragostini et al, 2006). This high TBP positivity rate associated to normal blood values highlighted the tolerance of the native sheep towards TBP infection and accounted for endemic TBD.

#### **3.1 Tick borne diseases in Apulian native sheep: A low income disease**

According to Townsend & Thirtle (2001), studies of the rates of return to research have usually been based on the implicit assumption that if there were no research, then there would be neither growth nor decline in output or productivity. In the case of livestock, particularly in those areas characterized by a sub-tropical disease ecology, the assumption is especially unreasonable. It ignores the losses that would have occurred in the absence of livestock health research, resulting in an underestimation of the rates of return. The financial impact of a range of clinical and subclinical diseases and mortalities on farms is difficult to assess because there are insufficient accurate survey data on their prevalence causes or production losses on a national basis. Thus demonstration of the economic advantage of animal health is one of the relevant issues in animal production. Pieragostini et al. (1996) carried out a four year study to check the economic and zoonotic importance of TBD on sheep farms.

To this purpose sheep belonging to breeds tolerant to TBD systematically underwent one prophylactic treatment with diminazene aceturate (Berenil, Hoechst, AG, Germany) in full spring before the mating season. Table 3 shows the results obtained. The comparison between the reproductive values in the treated sheep and in an untreated control group highlighted significant differences in fertility and fecundity, with the group of treated sheep that were more fertile and fecund. Pyroplasmosis, even though unapparent, represents an important cause of perturbation of animal welfare. The authors estimated relevant (30%) economic losses in non treated animals, thus defining pyroplasmosis as a "low income disease".

Tolerance to Tick-Borne Diseases in Sheep:

Modified from Pieragostini & Petazzi (1999).

high (Fig. 2).

identified in the blood smears.

Highlights of a Twenty-Year Experience in a Mediterranean Environment 457

Fig. 2. Graphical representation obtained by processing morbidity and mortality data evidencing the influence of genotype on tolerance to pyroplasmosis in sheep living in Apulia. 1, Apulian native breeds (Altamurana, Gentile di Puglia and Leccese);

2, Italian islander breeds (Comisana and Sardinian); 3, Crossbreds (Finnish x Altamurana, Finnish x Leccese, Friesian x Altamurana, Friesian x Leccese; Romanov x Altamurana, Romanov x Leccese); 4, Northern pure breeds (Finnish, Friesian and Romanov).

A further element to consider is that attempts to improve the productivity of Apulian breeds by crossing them with the above exotic breeds failed because of the high mortality in generations F1 and F2, almost solely due to TBD. Though the mortality rates in crossbred animals were lower than those registered in the respective parental pure breeds, the number of individuals killed by the impact with endoerythrocytic pathogens was in any case too

Pathogens were not accurately classified since the study analyzed data from farm records in which the veterinarians' diagnosis at death, due to TBD, always mentioned pyroplasmosis. The cases, which we were able to observe, concerned five Romanov sheep and seven Suffolk (occasionally found in the course of time and seriously ill prior to our visit). Examination of the animals always revealed classic symptoms of babesiosis and this was confirmed once the blood samples taken at the same time were analyzed. The haematological situation showed severe microcytic and hypochromic anaemia and *Babesia ovis (B. ovis)* was consistently

By contrast, among the resilient breeds of sheep, the animals infected with pyroplasmosis showed only a state of discomfort which usually does not last more than few days and is characterized by a brief rise in temperature, slight dejection in the form of a tendency to move away from the flock, loss of appetite which might also be very transitory, translucent

mucosae, slightly blueish against a pale background and in a few cases subicteric.


Table 3. Least-square means of the reproductive parameters in Altamurana and Leccese sheep and in the whole sample (Total), as a function of the prophylactic treatment against pyroplasmosis with diminazene aceturate (T = treated, NT = not treated). Modified from Pieragostini et al. (1996). Means within rows with different letters significantly differ: capital letters: P< 0.001; small letters: P<0.05.

#### **3.2 Breed sheep and TBD**

It is now generally acknowledged that importing exotic breeds can result in activities within the livestock sector that are uneconomic and/or have a negative impact on the environment. In many cases these activities are subsidized or otherwise provided for by development programs such as the case of Apulia which was documented in a study assessing attempts to introduce highly productive north European sheep breeds to Apulia (Pieragostini & Petazzi, 1999). The investigation analyzed data concerning the incidence and severity of pyroplasmosis in the five years spanning 1980-1984 on an experimental farm situated on the Murgia uplands in the province of Bari. The farm contained sheep belonging to gene pools of different geographical origin (Apulian, Italian island and north European breeds) or genotype classification (pure breeds or crossbreds) (Fig. 1).

The northern Finnish, Friesian and Romanov breeds were very susceptible to the disease; conversely, the native Apulian breeds showed very low rates of morbidity and mortality, followed in turn by breeds like Sardinian and Comisana, whose native areas have climatic and pedological characteristics similar to those of Apulia (Fig. 2). It is also worth noting that while the native and island breeds were regularly taken out to graze, the north European breeds were kept constantly under cover to reduce the likelihood of encountering ticks.

Fertility (%) 93a 65b 86a 62b 91A 64B Prolificacy (%) 139 131 144 142 138 134 Fecundity (%) 132a 89b 119a 90b 128A 89B

Table 3. Least-square means of the reproductive parameters in Altamurana and Leccese sheep and in the whole sample (Total), as a function of the prophylactic treatment against pyroplasmosis with diminazene aceturate (T = treated, NT = not treated). Modified from Pieragostini et al. (1996). Means within rows with different letters significantly differ: capital

It is now generally acknowledged that importing exotic breeds can result in activities within the livestock sector that are uneconomic and/or have a negative impact on the environment. In many cases these activities are subsidized or otherwise provided for by development programs such as the case of Apulia which was documented in a study assessing attempts to introduce highly productive north European sheep breeds to Apulia (Pieragostini & Petazzi, 1999). The investigation analyzed data concerning the incidence and severity of pyroplasmosis in the five years spanning 1980-1984 on an experimental farm situated on the Murgia uplands in the province of Bari. The farm contained sheep belonging to gene pools of different geographical origin (Apulian, Italian island and north European breeds) or

The northern Finnish, Friesian and Romanov breeds were very susceptible to the disease; conversely, the native Apulian breeds showed very low rates of morbidity and mortality, followed in turn by breeds like Sardinian and Comisana, whose native areas have climatic and pedological characteristics similar to those of Apulia (Fig. 2). It is also worth noting that while the native and island breeds were regularly taken out to graze, the north European breeds were kept constantly under cover to reduce the likelihood of

Fig. 1. Size of the investigated samples, clustered as sheep ecotypes. Modified from

NT (N = 259)

**Parameter** 

T (N =149)

letters: P< 0.001; small letters: P<0.05.

genotype classification (pure breeds or crossbreds) (Fig. 1).

**3.2 Breed sheep and TBD** 

encountering ticks.

Pieragostini & Petazzi (1999).

**Altamurana Leccese Total** 

NT (N = 89)

T (N = 198)

NT (N = 348)

T (N = 49)

Fig. 2. Graphical representation obtained by processing morbidity and mortality data evidencing the influence of genotype on tolerance to pyroplasmosis in sheep living in Apulia. 1, Apulian native breeds (Altamurana, Gentile di Puglia and Leccese); 2, Italian islander breeds (Comisana and Sardinian); 3, Crossbreds (Finnish x Altamurana, Finnish x Leccese, Friesian x Altamurana, Friesian x Leccese; Romanov x Altamurana, Romanov x Leccese); 4, Northern pure breeds (Finnish, Friesian and Romanov). Modified from Pieragostini & Petazzi (1999).

A further element to consider is that attempts to improve the productivity of Apulian breeds by crossing them with the above exotic breeds failed because of the high mortality in generations F1 and F2, almost solely due to TBD. Though the mortality rates in crossbred animals were lower than those registered in the respective parental pure breeds, the number of individuals killed by the impact with endoerythrocytic pathogens was in any case too high (Fig. 2).

Pathogens were not accurately classified since the study analyzed data from farm records in which the veterinarians' diagnosis at death, due to TBD, always mentioned pyroplasmosis. The cases, which we were able to observe, concerned five Romanov sheep and seven Suffolk (occasionally found in the course of time and seriously ill prior to our visit). Examination of the animals always revealed classic symptoms of babesiosis and this was confirmed once the blood samples taken at the same time were analyzed. The haematological situation showed severe microcytic and hypochromic anaemia and *Babesia ovis (B. ovis)* was consistently identified in the blood smears.

By contrast, among the resilient breeds of sheep, the animals infected with pyroplasmosis showed only a state of discomfort which usually does not last more than few days and is characterized by a brief rise in temperature, slight dejection in the form of a tendency to move away from the flock, loss of appetite which might also be very transitory, translucent mucosae, slightly blueish against a pale background and in a few cases subicteric.

Tolerance to Tick-Borne Diseases in Sheep:

\*P<0.05;\*\*P<0.01; n.s.= not significant.

where endoerythrocytic parasites are endemic.

**3.4 Anaplasmosis in naturally infected splenectomized sheep** 

Highlights of a Twenty-Year Experience in a Mediterranean Environment 459

**Haematological parameters Samples Significance** 

Anaplasmosis is one of the most important tick-borne diseases of ruminants worldwide. The disease is caused by infection of animals with the obligate intraerythrocytic bacteria *Anaplasma* spp. which is classified in the family *Anaplasmataceae*, order *Rickettsiales* (Dumler et al, 2001). This section includes some experiences with sheep splenectomy and describes disease onset and course in eight splenectomized TBD-tolerant sheep that were naturally infected with piroplasms. Though the trials had been performed in different time periods, the results obtained were very similar and the facts surrounding the experiments gave us both general and specific insights into the field of splenectomy of carrier sheep from areas

Particularly in the first trial, the surgical operation had two purposes: a) to evaluate the rôle of the spleen as a filter-pad to check parasites and as modulator of the direct response to anemia;

The following trials were mainly related to the need to obtain *A. ovis* which was isolated

Splenectomy was slightly traumatic for all the subjects and 24 hours after the surgical operation the sheep showed normal functions. The sheep were identified with female names for easier checking. Clinical evaluation was done on a daily basis and rectal temperatures were recorded every morning for 12 weeks post splenectomy. Blood and serum samples were routinely collected twice a week during the observation period. Haematological variables were evaluated using a haematology analyzer. The erythrocyte fragility test was performed by exposing erythrocytes to hypotonic saline solutions decreasing by 0.02% starting from 0.86%. Parasites in the blood were checked by Giemsa staining every 3 days. During the acute phase of the disease, the most important haematological values, erythrocyte fragility and parasitaemia were monitored daily. In the case of Gilda, Lina and Zoppina, which were part of the experiment to check the response to *A. ovis* infection of

b) to obtain a high number of parasites in the blood to prepare a local specific antigen.

from splenectomized sheep allowed to be naturally infected pasturing in tick areas.

RBC (106/l) 9.50.19 6.30.70 \*\* Hb (g/dl) 9.90.18 9.20.50 \* PCV (g/dl) 32.30.55 30.30.50 \*\* MCV (fl) 34.31.43 48.71.29 \*\* MCH (pg) 10.40.46 15.20.42 \*\* MCHC (g/dl) 30.50.44 31.20.40 n.s. WBC (103/l) 9.60.40 9.50.37 n.s. MCF (g%NaCl for 50% haemolysis) 0.720.02 0.820.03 \*\* Table 4. Comparison of the haematological parameters recorded in healthy and affected Altamurana sheep (mean values standard errors). Modified from Pieragostini & Petazzi, 1999. (RBC=Red Blood Cells; Hb=Haemoglobin %; PCV= Packed Cell Volume; MCV= Mean Corpuscular Volume; MCH=Mean Corpuscular Haemoglobin, MCHC=Mean Corpuscular Haemoglobin Content; WBC=White Blood Cells; MCF=Mean Corpuscolar Fragility)

Healthy (N =22) Affected (N =28)
