PUBLIC HEALTH HAZARD ASSOCIATED WITH CONSUMPTION OF MILK FROM CATTLE INFECTED WITH SUBCLINICAL MASTITIS IN ASSIUT GOVERNORATE

Dept. of Food Hygiene,

Fac. of Vet. Med., Assiut Univ.

PUBLIC HEALTH HAZARD ASSOCIATED WITH CONSUMPTION OF MILK FROM CATTLE INFECTED WITH SUBCLINICAL MASTITIS IN ASSIUT GOVERNORATE

(With 6 Tables and 4 Figures)

By

T. EL-BASSIONY; ENAS EL-PRINCE;

AMAL A. ABDEL-HALEEM* and O.A. SADEK*

* Animal Health Research Institute

(Received at 16/4/2009)

المخاطر الصحية على الإنسان المرتبطة بتناول ألبان الماشية المصابة

بالتهاب الضرع الغير ظاهري بمحافظة أسيوط

توفيق البسيوني ، إيناس البرنس ، آمال على عبد الحليم ، أنسى صادق

يعتبر التهاب الضرع تحت الأکلينيکى من الأمراض المؤثرة اقتصاديا وصحيا لما يسببه من نقص في إنتاج اللبن بالإضافة إلى نقل بعض الميکروبات الخطيرة للإنسان. نظرا لأهمية هذا المرض فقد تم تجميع عدد 787 عينة من ألبان الأبقار والجاموس, وتم فحصها بکتريولوجيا وميکولوجيا لعزل وتصنيف المسبب. بفحص 379 عينة من لبن الأبقار باختبارات CMT  و  MWST، کانت النتائج ايجابية بنسبة 29.55 و 30.87%. وبفحص 408 عينة من لبن الجاموس کانت النتائج ايجابية بنسبة 11.76 و 12.01% لکلا الاختبارين على الترتيب. وقد دل الفحص البکتريولوجى على أن النتائج کانت ايجابية بنسب 28.50 و 10.54% لکل من الأبقار والجاموس على الترتيب. أما فى حالة نتائج  الفحص الميکولوجي فقد کانت ايجابية بنسب 16.09 و 0.25% على الترتيب. کما تبين أن نسبة عزل البکتريا المسببة لإلتهاب الضرع تحت الاکلينيکي في الأبقار هي: 15.57، 9.50، 2.64، 1.32، 4.49، 13.19، 1.32، 1.58 و 0.26% لکل من المکور العنقودي الذهبي، المکور العنقودي السالب التجلط، المکور السبحي بيوجين، المکور السبحي اجالاکتيا، المکور السبحي ديس جالاکتيا، الميکروب القولوني کلبسيلا نوموني، ستروباکتر داي فيرساس وبروتياس ميرابيليز. بينما في الجاموس کانت النسب کالتالي: 8.33، 1.96، 0.25، 0.25 و 1.72% لکل من المکور العنقودي الذهبي، المکور العنقودي السالب التجلط، المکور السبحي بيوجين، المکور السبحي ديس جالاکتيا والکورايني باکتريام بوفس على الترتيب. أما الخمائر والفطريات فکانت نسب عزلها في الأبقار کالتالي: 4.75، 1.58، 2.38، 2.90، 1.32، 1.06، 0.53، 0.79، 0.26، 0.26،0.26، 0.53، 0.26، 0.26، 0.26، 0.26و0.26% کل من

 Candida albicans,  Candida tropicalis,  Candida krusei,  Candida sp. , Geotrichum candidum,  Aspergillus niger,  Cladosporium cladosporioides, Fusarium proliferatum,  Penicillium duclauxi,  Rhodotorula sp., Stachybotrys elegans,  near to Pyssochlamys nivea,  Alternaria alternate,  Stemphylium botryosum,  Thermoascus aurantiacus,  Trichosporon cuteanum and sterile myceliumعلى الترتيب .

بينما تم عزل Phialophora sp. من الجاموس بنسبه 0.25%. هذا وقد تمت مناقشة الأهمية الصحية والوبائية والاقتصادية لمرض التهاب الضرع تحت الاکلينيکي في الإنسان والحيوان والشروط الواجب إتباعها لمنع انتشاره واختباره في مزارع الألبان المختلفة لدرء خطره على الإنسان.

SUMMARY

Seven hundred and eighty seven random milk samples were collected from cows and buffaloes at different localities and farms in Assiut Governorate. These samples represented by 379 and 408 quarter milk samples from 105 cows and 105 buffaloes, respectively. Animal-wise incidence of Subclinical Mastitis (SCM) based on California Mastitis Test (CMT) and Modified Whiteside Test (MWST) were 59.05 & 60.95% positive in cows and 33.33% & 33.33% positive in buffaloes, respectively. The animal-wise incidences of SCM in cows & buffaloes by bacteriological and mycological examinations were 60.95 & 28.57% and 34.29 & 0.95% positive, respectively. The most common bacteria causing SCM in cows were Staph. aureus, coagulase negative staphylococci (CNS), Str. pyogenes, Str. agalactiae, Str. dysgalactiae, E. coli, Klebsiella pneumoniae, Citrobacter diversus and Proteus mirabilis, respectively. While, in buffaloes were Staph. aureus, CNS, Str. pyogenes, Str. dysgalactiae and Corynebacterium bovis. Additionally, the most common yeasts and molds causing SCM in cows were Candida albicans, Candida tropicalis, Candida krusei, Candida sp., Geotrichum candidum, Aspergillus niger, Cladosporium cladosporioides, Fusarium proliferatum, Penicillium duclauxi, Rhodotorula sp., Stachybotrys elegans, near to Pyssochlamys nivea, Alternaria alternata, Stemphylium botryosum, Thermoascus aurantiacus, Trichosporon cuteanum and sterile mycelium. However, Phialophora sp. was only isolated from buffaloes in a percentage of 0.25%.

Key words: Subclinical mastitis; cow's, buffalo's milk; public health hazards

INTRODUCTION

Mastitis is an inflammation of udder, usually as a result of microbial infection by invading bacteria or other microorganisms including fungi and possibly viruses. Mastitis can generally be characterized as clinical, subclinical and chronic disease (Watts, 1990). The economic impact of both clinical and subclinical forms of mastitis is large in the current dairy industry. Furthermore, in dairy industry, SCM remains an important cause of reduced milk production and poor quality value products such as cheese or casein (Roux et al., 1995). Losses occur from decreased milk production, treatment and labour costs, non deliverable milk and veterinary fees, reduced milk quality, reduced milk price, increased risk of subsequent mastitis and increased risk of culling or death of the animal (Harmon, 1994). Diagnosis of acute mastitis presents little difficulty as compared to subclinical by ordinary clinical tests and as a result, SCM is responsible for great losses to the dairy industry therefore, more attention has been given for the diagnosis of SCM by indirect tests (Joshi et al., 1976). It is the most serious form as both infected udder and milk show no obvious clinical abnormalities, whereas, several causative organisms are discharged with the milk for long time. Regarding public health importance, mastitis is considered of quite vital importance due to its association with many zoonotic diseases in which milk act as a vehicle of infection. Of the various conditions of udder, the SCM is attracting the eye world over as this condition not only leads to suboptimal milk production but it also results in transmission of certain diseases to human beings too (Tijare et al., 2000).As a results of the economic and public health importance of SCM, the objective of this work was done to detect SCM in cows and buffaloes by using screening or field tests, bacteriological examination as well as mycological examination.

Materials and Methods

Milk samples:

A total number of 787 milk samples were collected from apparently healthy cows and buffaloes at different farms in Assiut Governorate, comprising 379 quarter milk samples from 105 lactating cows and 408 quarters of milk samples from 105 lactating buffaloes.

I-Screening tests to detect SCM:

A- CMT according to Schalm et al. (1971).

B- MWST according to Murphy and Hanson (1941).

II- Microbiological examination of subclinical mastitic milk samples:

A- Isolation of members of Family Enterobacteriaceae according to Cruickshank et al. (1975); Finegold and Martin (1982); and Koneman et al. (1992).

B- Isolation of Staph. aureus (Bailey and Scott, 1994) .

C- Isolation and identification of Streptococci (Cruickshank et al., 1975 and Quinn et al., 1994).

D- Isolation and identification of yeasts and molds

Fungal cultures were kindly identified by staff members of the Assiut University Mycological Centre (AUMC), Assiut, Egypt, using the following references: Kwon-Chung and Bennett (1992), Moubasher (1993), Odds and Bernaerts (1994) and Hoog et al. (2000)

RESULTS

Table 1: Quarter-wise incidence of SCM in cow's and buffalo's milk samples based on the results of CMT and MWST.

Table 2: Animal-wise incidence of SCM in cow's and buffalo's milk samples based on the results of CMT and MWST.

Table 3: Quarter-wise incidence of SCM in cow's and buffalo's milk samples based on the results of bacteriological and mycological examination.

Table 4: Animal-wise incidence of SCM in cow's and buffalo's milk samples based on the results of bacteriological and mycological examination.

Table 5: Incidence of the isolated bacteria causing SCM in the examined cow's and buffalo's milk samples.

Table 6: Incidence of the isolated yeasts and molds causing SCM in cow's and buffalo's milk samples.

Fig. 1: Incidence of the isolated bacteria causing SCM in the examined cow's and buffalo's milk samples.

Fig. 2: Incidence of the isolated yeasts and molds causing SCM in cow's and buffalo's milk samples.

Fig. 3: Incidence of the isolated yeasts causing subclinical mastitis in cow's milk samples

Fig. 4: Incidence of the isolated molds causing subclinical mastitis in cow's milk samples

Discussion

Data presented in Table 1, showed the quarter-wise incidence of SCM in cow's and buffalo's milk samples based on the results of both CMTand MWST. Out of 379 quarters cow's milk, 112 (29.55%) samples were positive for CMT. Somewhat similar results were recorded by Hatem et al., (1984) (28.28%). Lower findings were estimated by Singh et al., (1982) (12.33%) and Saini et al., (1994) (4.87%), however, higher incidence was detected by Mohamed et al., (1993) (46%). Out of 408 quarters buffalo's milk examined by CMT, 48 (11.76%) were positive (Table 1). Nearly similar results were recorded by Singh et al., (1982) (9.54%) and Naiknaware et al., (1998) (9.87%). On the contrary, a lower level was recorded by Saini et al., (1994) (2.59%), however, relatively higher results were reported by El-Balkemy et al., (1997) (26.34%) and Salama, (2004) (43.13%).

Regarding quarter-wise incidence of SCM in cow's and buffalo's milk samples based on the result of MWST, 117 samples (30.87%) of cow's milk were positive to MWST. This incidence was in accordance with that estimated by Tijare et al. (1999) (28.69%). Singh et al. (1982) and El-Balkemy et al. (1997) recorded lower findings of 10.73 and 17.47%, respectively. In contrast, higher result was incriminated by Wahba et al. (2005) (93.8%). Concerning buffalo's quarter milk, 49 samples (12.01%) were positive to MWST (Table 1). Lower result was indicated by Singh et al. (1982) (7.14%), however, El-Balkemy et al. (1997) detected relatively higher incidence of 30.10%.

Table 2, estimated the animal-wise incidence of SCM in cow's and buffalo's milk samples based on the results of both CMT and MWST. Out of 105 cows examined, 62 animals (59.05%) gave positive result, which in harmony with that estimated by Tijare et al. (1999) (57.98%), while, lower percentages were recorded by Saini et al. (1994) (17.33%), Singh et al. (1994) (49.29%), El-Balkemy et al. (1997) (31.98%), and Mukherjee and Dash (2003) (52.32%). However, higher finding was recorded by Prasad et al. (2001) (61.32%). Concerning buffaloes, 35 animals (33.33%) were positive to CMT which go parallel with that estimated by Farah and Kaldes (1999) (36.6%). Saini et al. (1994) and Naiknaware et al. (1998) recorded lower incidences (9.59 and 28.63%, respectively), however, El-Balkemy et al. (1997) showed higher percentages of 42.55%.

The animal-wise incidences based on the results of MWST, showed that out of 105 cows, 64 animals (60.95%) were positive (Table 2). Lower results were estimated by El-Balkemy et al. (1997) (34.26%), however somewhat higher result was detected by Tijare et al. (1999) (70.59%). In case of buffalo's milk samples, 35 animals (33.33%) were positive. Higher incidences were estimated by El-Balkemy et al. (1997) (48.93%), and Farah and Kaldes (1999) (63.3%).

The quarter-wise incidence of SCM in cow's and buffalo's milk samples based on the results of bacteriological and mycological examination was illustrated in Table 3. Out of 379 quarters cows milk samples examined, 108 (28.50%) were positive. The results were in agreement with that evaluated by Pardo et al. (1998) (26.5%). In the contrary, Saini et al. (1994) recorded relatively higher incidence of 76.13%. Moreover, in case of buffalo's milk samples, 43 (10.54%) were positive and 365 (89.46%) were negative. Higher percentages were estimated by Tijare et al. (1999) (26.77%). Regarding the results of mycological examination, 61 (16.09%) and one (0.25%) milk samples of cow's and buffalo's were positive, respectively (Table 3).

The obtained results in Table 4 revealed that, the animal-wise incidence of SCM in cows was 60.95% bacteriologically positive. Extremely lower results were stated by Singh et al. (1982) (18.4%) and Petrović et al. (1997) (51.0%), whereas, Abdel-Ghani (2005) recorded higher result (67.5%). In buffalo's milk, 30 (28.57%) samples were positive. Singh et al. (1982) revealed a lower incidence of 13.2%, while, Tijare et al. (1999) estimated extremely higher incidence of 67.22%. Furthermore, mycologically 34.29 and 0.95% were positive for cows and buffaloes, respectively.

The incidences of isolated bacteria causing SCM in the examined cow's milk samples were showed in Table 5 and Fig. 1. 59 isolates (15.57%) of Staph. aureus were recovered from the examined samples. This result was in accordance with that obtained by Kotb (2006) (14%). Lower results were obtained by Petrović et al. (1997) (4.69%), Pardo    et al. (1998) (8.52%), Wahba et al. (2005) (10%) and Abdel-Hameed (2006) (9.28%). However, higher incidences were recorded by Singh    et al. (1994) (35.38%), Mokhbatly et al. (2001) (40.90%), Al-Hawary   et al. (2003) (29.2%) and Abdel-Ghani (2005) (25.6%). Also, 36 isolates (9.50%) of CNS were obtained which relatively similar to the result recorded by Singh et al. (1994) (10%) however, lower incidence (5.4%) was stated by Abdel-Hameed (2006). Extremely higher percentages were obtained by Pardo et al. (1998) (64.2%), Abdel-Ghani (2005) (34.1%) and Wahba et al. (2005) (66%). In addition, 10 isolates (2.64%) of Strept. pyogenes were described in Table 5 and Fig. 1. Five isolates (1.32%) of Strept. agalactiae were also isolated which in accordance with that estimated by Al-Hawary et al. (2003) (1.5%), while, lower results were reported by Petrović et al. (1997) (0.34%) and Abdel-Hameed (2006) (0.7%). On the other hand, Singh et al. (1994) and Mokhbatly et al. (2001) recorded higher incidences of 10 and 11.36%, respectively. Concerning Strept. dysgalactiae, 17 isolates (4.49%) were recovered from the examined samples which were in harmony with that estimated by Mokhbatly et al. (2001) (4.55%). Singh et al. (1994) and Abdel-Hameed (2006) stated slightly lower incidences of 3.08 and 3.88%, respectively. Moreover, 50 isolates (13.19%) of E. coli were recorded and this result was nearly similar to that postulated by Abdel-Ghani (2005) (12.8%). In contrary, lower results were incriminated by Singh et al. (1994) (5.38%) and Al-Hawary et al. (2003) (6.2%). While, extremely lower incidences were obtained by Pardo et al. (1998) (2.84%), and Abdel-Hameed (2006) (1.66%). However, Mokhbatly et al. (2001) and Kotb (2006) stated higher incidences of 18.20 and 15.0%, respectively. With regard to Klebsiella pneumoniae 5 isolates (1.32%) were obtained which was somewhat lower than that estimated by Mokhbatly et al. (2001) (2.27%) and Kotb(2006) (3%). Six isolates (1.58%) of Citrobacter diversus were obtained, while, only one isolate (0.26%) of Proteus mirabilis was recovered from tested samples and Abdel-Ghani (2005) reported a higher incidence of 0.9%.

In case of buffalo's milk, it is clear that, 34 isolates (8.33%) of Staph aureus were recovered from the examined samples. Lower results were estimated by Tijare et al. (1999) (2.52%), however, higher incidences with a great extent were stated by El-Balkemy et al. (1997) (35.41%) and Mokhbatly et al. (2001) (26.67%). Also, 8 isolates (1.96) of CNS were obtained which is extremely lower than that stated by Tijare et al. (1999) (36.48%). In addition, one isolate (0.25%) of each Strept. pyogenes and Strept. dysgalactiae was isolated. El-Balkemy et al. (1997) (2.08%) and Mokhbatly et al. (2001) (4.44%) recorded higher incidences of a great extent. Concerning Corynebacterium bovis, 7 isolates (1.72%) were recovered which is lower than the results stated by Tijare et al. (1999)(6.29%) and Mokhbatly et al. (2001) (2.22%).

It is evident that, Staph. aureus was the highest isolated microorganism from both cow's and buffalo's milk. This may be due to its ubiquitous nature and due to indiscriminate using of antibiotics resulting in the emergence of resistant strains (Singh et al., 1994). Also, skin of milkers hand may play a role in its transmission from teat to teat and from animal to animal. Moreover, it is evident that, Strept. agalactiae couldn't be isolated from buffalo's milk. 

The incidences of the isolated yeasts and molds causing SCM in cow's milk samples were shown in Table 6 and Fig. 3 & 4. 18 isolates (4.75%) of Candida albicans were recovered from the examined samples, Ahmed (1981) stated higher incidence of 60%. Six strains (1.58%) of Candida tropicalis were isolated and this finding is lower with a great extent than that recorded by Ahmed (1981) (22.5%) in Friesian cattle. Also, 9 isolates (2.38%) of Candida krusei were obtained which is lower than that estimated by Ahmed (1981) (5%). In addition, 11 isolates (2.90%) of other Candida sp. were recovered from the examined samples. With regard to Geotrichum candidum, 5 isolates (1.32%) were obtained which is lower than that reported by Ahmed (1981) (5%). Concerning molds, 4 isolates (1.06%) of Aspergillus niger were isolated. Higher results were estimated by Ahmed (1981) (6.67%) and Abdel-Ghani (2005) (5.9%). Also, Cladosporium cladosporioides, Fusarium proliferatum, Penicillium duclauxi, Rhodotorula sp., Stachybotrys elegans, near to Pyssochlamys nivea, Alternaria alternata, Stemphylium botryosum, Thermoascus aurantiacus, Trichosporon cuteanum and sterile mycelium were isolated in percentages of 0.53, 0.79, 0.26, 0.26, 0.26, 0.53, 0.26, 0.26, 0.26, 0.26 and 0.26%, respectively.

Only one isolate (0.25%) of Phialophora sp. was isolated from buffalo's milk samples (Table 6). It is clear that, no yeasts or other molds could be isolated from buffalo's milk and this could be attributed to the structure of buffalo's teat canal or may be due to the genetic character of buffaloes which render it more resistant to mycotic infection or other factors which need further investigations.

Under the condition of this investigation, it can be concluded that, CMT and MWST were highly sensitive for detection of SCM and their application lead to earlier detection of infected animal and its isolation either for culling or therapy. Buffaloes were less susceptible to SCM as compared to cow. Staphylococci either Staph. aureus or CNS were the most causative agents causing SCM followed by E. coli. In cows, yeasts were higher than molds for causing SCM however, buffaloes somewhat resist to mycotic SCM.

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